See discussions, stats, and author profiles for this publication at: https://www.researchgate.

net/publication/11328388

The La protein

Article in Annual Review of Biochemistry · February 2002

DOI: 10.1146/annurev.biochem.71.090501.150003 · Source: PubMed

CITATIONS READS

354 4,617

2 authors, including:

Tommy Cedervall
Lund University
78 PUBLICATIONS 12,518 CITATIONS

SEE PROFILE

All content following this page was uploaded by Tommy Cedervall on 19 May 2014.

The user has requested enhancement of the downloaded file.

 Annu. Rev. Biochem. 2002. 71:375– 403
DOI: 10.1146/annurev.biochem.
Copyright © 2002 by Annual Reviews. All rights reserved

THE LA PROTEIN
Sandra L. Wolin and Tommy Cedervall
Departments of Cell Biology and Molecular Biophysics and Biochemistry, Howard
Hughes Medical Institute, Yale University School of Medicine, 295 Congress Avenue,
New Haven, Connecticut 06536; e-mail: sandra.wolin@yale.edu; tcedervall@yahoo.com

Key Words RNA polymerase III, tRNA maturation, small RNAs, nuclear
retention, RNA binding
f Abstract Ubiquitous in eukaryotic cells, the La protein associates with the 3⬘
termini of many newly synthesized small RNAs. RNAs bound by the La protein
include all nascent transcripts made by RNA polymerase III as well as certain small
RNAs synthesized by other RNA polymerases. Recent genetic and biochemical
analyses have revealed that binding by the La protein protects the 3⬘ ends of these
RNAs from exonucleases. This La-mediated stabilization is required for the normal
pathway of pre-tRNA maturation, facilitates assembly of small RNAs into functional
RNA-protein complexes, and contributes to nuclear retention of certain small RNAs.
Studies of mutant La proteins have given some insights into how the La protein
specifically recognizes its RNA targets. However, many questions remain regarding
the molecular mechanisms by which La protein binding influences multiple steps in
small RNA biogenesis. This review focuses on the roles of the La protein in small
RNA biogenesis and also discusses data that implicate the La protein in the
translation of specific mRNAs.

CONTENTS
INTRODUCTION . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 376
ABUNDANCE AND SUBCELLULAR LOCATION. . . . . . . . . . . . . . . . . . . 377
STRUCTURAL FEATURES OF LA PROTEINS. . . . . . . . . . . . . . . . . . . . . 378
Domain Organization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 378
The Family of La Motif–Containing Proteins . . . . . . . . . . . . . . . . . . . . . . 381
FUNCTIONS OF THE LA PROTEIN. . . . . . . . . . . . . . . . . . . . . . . . . . . . 384
La Is Required for the Normal Pathway of Pre-tRNA Maturation . . . . . . . . . . 384
La Stabilizes Other Nascent RNAs from Exonuclease Digestion . . . . . . . . . . 386
La May Contribute to Nuclear Retention of Nascent Transcripts . . . . . . . . . . 388
Is La a Transcription Factor for RNA Polymerase III? . . . . . . . . . . . . . . . . 390
Possible Roles for La in mRNA Translation. . . . . . . . . . . . . . . . . . . . . . . 391
Other Potential Roles for the La Protein . . . . . . . . . . . . . . . . . . . . . . . . . 394
RNA BINDING BY THE LA PROTEIN . . . . . . . . . . . . . . . . . . . . . . . . . . 394
RNA Sequences Recognized by La . . . . . . . . . . . . . . . . . . . . . . . . . . . . 394
Features of La That Contribute to RNA Recognition . . . . . . . . . . . . . . . . . 395

0066-4154/02/0707-0375$14.00 375
376 WOLIN y CEDERVALL

Is Oligomerization Required for La Function? . . . . . . . . . . . . . . . . . . . . . 397

PERSPECTIVES AND FUTURE DIRECTIONS . . . . . . . . . . . . . . . . . . . . . 398

INTRODUCTION

A highly abundant nuclear phosphoprotein known as the La protein is the first

protein that associates with all newly synthesized RNA polymerase III tran-
scripts. The La protein (also called SS-B) was first described as an autoantigen
in patients suffering from the rheumatic diseases systemic lupus erythematosus
and Sjogren’s syndrome (1, 2). (The name La derives from the name of the
patient in which the antibody was detected, and SS-B refers to the prevalence of
the antibodies in Sjogren’s syndrome.) Although La was first characterized as a
human protein, homologs have been identified in a wide variety of eukaryotes
(3–12). Experiments in which patient anti-La antibodies were used to immuno-
precipitate La ribonucleoproteins (RNPs) from radiolabeled cell extracts revealed
that the La protein associated with a very large number of nascent small RNAs.
RNAs identified in anti-La immunoprecipitates include pre-tRNAs, pre-5S
rRNA, U6 small nuclear RNA (snRNA), RNase P RNA, MRP RNA, 7SL RNA,
Y RNAs, rodent 4.5SI and 4.5SII RNAs, and transcripts of Alu sequences
(13–20). Experiments using virus-infected cells revealed that La also binds a
number of viral-encoded RNAs, including the adenovirus-encoded VA RNAI
and VA RNAII (13, 21), the Epstein-Barr virus– encoded EBER 1 and EBER 2
RNAs (22), and the leader RNAs of several negative-strand viruses (23–26).
For the majority of cellular RNAs, La binds the precursors, rather than the
mature RNAs. For example, the U6 RNAs bound by the human La protein are
extended at the 3⬘ end and contain fewer modifications than the mature RNA
(20). Similarly, the La protein binds precursors to tRNAs and 5S rRNA, but does
not bind the corresponding mature RNAs (15). The reason that the La protein
associates primarily with nascent RNA polymerase III transcripts became clear
when it was shown that at least part of the recognition site for the protein is
UUUOH, which is at the 3⬘ end of virtually all newly synthesized RNA
polymerase III transcripts (27–29). These terminal uridylates are usually
removed during RNA maturation, thus eliminating the binding site for the La
protein. Consistent with this, the yeast La protein also binds certain nascent RNA
polymerase II–transcribed small RNAs that end in UUUOH (30, 31).
Because the majority of RNAs associated with the La protein are newly
synthesized RNA polymerase III transcripts, the protein has long been assumed
to function in some aspect of the biogenesis of these RNAs. One of the first
functions proposed for La was as a transcription termination factor for RNA
polymerase III (32–38). However, recent experiments have revealed that tran-
scription of several RNA polymerase III genes is efficient in both yeast and
vertebrate cell extracts lacking detectable La protein (39 – 43). More recently, it
has become clear that a major role of La is to stabilize newly synthesized small
 THE LA PROTEIN 377

RNAs from exonuclease digestion (30, 31, 34, 39 – 41, 44, 45). This La-mediated
stabilization of 3⬘ ends has different consequences for the various RNAs bound.
For pre-tRNAs, binding by La is required for the normal endonucleolytic
removal of the 3⬘ trailer. In the absence of La, the 3⬘ trailer is removed by
exonuclease(s) (10, 39, 44, 46). For certain other small RNAs, binding by La
contributes to nuclear retention of the nascent transcripts (47– 49) and/or assem-
bly of the RNAs into functional RNPs (30, 40). Because transient binding by the
La protein facilitates the correct fate of many newly synthesized small RNAs, La
has been proposed to function as a molecular chaperone for small RNA
biogenesis (30, 40, 47, 50).
In this review, we focus primarily on the roles of the La protein in small RNA
biogenesis. Although a high-resolution structure of La is not yet available,
studies of mutated and truncated La proteins have given some insights into the
features of La that are required for function. Other potential roles for the La
protein are also discussed, most notably experiments that implicate this normally
nuclear protein in facilitating translation initiation on certain viral and cellular
mRNAs.

ABUNDANCE AND SUBCELLULAR LOCATION

The La protein is an exceedingly abundant protein. The human protein has

been estimated to be present in 2 ⫻ 107 copies per cell (32), making it about
as abundant as a ribosomal protein. Quantitation of the amount of La protein
present in human and yeast extracts has resulted in estimates of 50 nM for a
human S100 extract (44) and 12 nM for a Saccharomyces cerevisiae S100
extract (39).
Immunofluorescence experiments have revealed that in virtually all cell types
and species, La is largely confined to the nucleus (7, 11, 14, 51–56). Within the
nucleus, La is both nucleoplasmic and nucleolar (14, 57–59). Despite this
localization, the La protein is found in the cytoplasm when cells are fractionated
using standard procedures (13, 29), indicating that the protein readily leaks from
the nucleus. However, La is almost exclusively found in nuclear fractions when
cells are separated into cytoplasts and karyoplasts under conditions that minimize
nuclear leakage (60). Also, when Xenopus oocytes are separated into nuclear and
cytoplasmic fractions by manual enucleation, the endogenous La protein is
primarily found in the nucleus (62).
Although La is largely nuclear under normal cellular conditions, a fraction
of the La protein becomes cytoplasmic in certain circumstances. During
poliovirus infection of mammalian cells, a portion of the La protein redis-
tributes to the cytoplasm (50). A possible mechanism was suggested by the
finding that a poliovirus-encoded protease cleaves La in the C terminus,
removing the nuclear localization signal (63). Similarly, the La protein
becomes cytoplasmic during apoptosis of mammalian cells (64 – 66), possibly
378 WOLIN y CEDERVALL

owing to removal of the nuclear localization signal by a caspase-like pro-

tease (65, 66). Because the distribution of the mammalian La protein changed
when cells were treated with certain metabolic inhibitors, the La protein
has been proposed to shuttle between the nucleus and cytoplasm (67).
However, using conventional heterokaryon assays for shuttling (68), others
have been unable to confirm that La is a shuttling protein (GJ Pruijn, personal
communication).

STRUCTURAL FEATURES OF LA PROTEINS

Domain Organization
Alignment of La proteins from species ranging from humans to trypanosomes
reveals that these proteins can be divided into at least three regions (Figure 1).
The N terminus of all known La proteins contains the ⬃60 –amino acid La motif
[previously called the La domain (10)], a highly conserved sequence that is also
present in a number of otherwise unrelated proteins (7, 56, 69, 70). Following the
La motif, there is a less well conserved RNA recognition motif (RRM; also
known as RNP domain) (71, 72) and a highly charged, weakly conserved C
terminus. Consistent with the idea that the C terminus represents a distinct
domain of La, this portion of the human protein can be severed from the
N-terminal half of the protein by a variety of proteases (4, 73). The molecular
size of the La protein varies from ⬃50 kilodaltons in vertebrates to 32 kilodaltons
in the yeast S. cerevisiae (Figure 1). Almost all of the additional mass of the
larger La proteins is due to an expanded C-terminal domain (Figure 1).
On the basis of sequence analyses, it has been proposed that the highly
conserved N terminus of La, consisting of the La motif and adjacent amino acids,
folds into an RNA recognition motif (74, 75). In this model, the N-terminal half
of La contains two RRMs (74, 75). However, other modelings of La protein
structure (56, 76) have failed to support the idea that the La motif folds into a
canonical RRM. Moreover, although there are two short conserved motifs, RNP1
and RNP2, within every RRM, it is largely the overall structure, rather than the
amino acid sequence, that is conserved between the RRMs of otherwise unrelated
proteins (77). In contrast, the La motifs of both bona fide La proteins and the
other La motif– containing proteins are highly related throughout the entire motif
(Figure 2). Thus, although the structure formed by the La motif will be uncertain
until high-resolution structural data are available, the fact that the entire motif is
present, essentially intact, in otherwise unrelated proteins suggests that it has
evolved to carry out a highly specific function.
The C-terminal domain is the least conserved part of the La protein, varying
in both size and sequence between species. This portion of the La protein
(defined as being C-terminal to the central RRM), usually contains between 40%
and 50% charged residues (7, 78). The lower degree of conservation of the
 THE LA PROTEIN 379

Figure 1 Structure of La proteins. The human, Drosophila, C. elegans, S.

pombe, S. cerevisiae, and T. brucei proteins are aligned. For each protein, the
length in amino acids is indicated on the right. All these proteins, with the
exception of C. elegans C44E4.4 (accession number AAB54169), have been
demonstrated to bind newly synthesized RNA polymerase III transcripts in vivo
(7, 10, 15; E Ullu & S Wolin, unpublished data) and are thus bona fide La
proteins. Of the three C. elegans La motif– containing proteins, only C44E4.4 is
homologous to authentic La proteins throughout its length, making it likely to be
the C. elegans La protein. A potential atypical RRM has been noted in the
C-terminal domain of vertebrate La proteins (79). Modeling of this portion of
the Drosophila and C. elegans proteins gives weak evidence for the presence
of the RRM in Drosophila and does not predict an RRM in the C. elegans pro-
tein (J Bruning & Y Shamoo, personal communication). The position of a
short basic motif (SBM) in the human protein (123) is indicated, as is a region of
the protein proposed to function in dimerization (81). Vertebrate La pro-
teins contain a potential Walker A motif, GXXXXXGKX, within the short
basic motif (5). Nuclear localization sequences (NLS) are indicated for the five
studied La proteins (11, 62, 73a). The S. cerevisiae NLS has not been finely
mapped, but has been located within a 113-amino-acid fragment that includes the
RRM (73a).

C-terminal portion is emphasized by the finding that almost all differences

between the human and mouse proteins are found in the C terminus (5). The
C-terminal region is quite variable in length, ranging from ⬃70 amino acids in
the yeasts S. cerevisiae and S. pombe to more than 220 amino acids in humans
and Xenopus laevis. At least some of the additional size of vertebrate La proteins
comes from a potential atypical RRM in this region (79). A possible Walker A
motif, a nucleoside triphosphate– binding sequence found in many ATP-binding
proteins (80), has also been noted in the C terminus of vertebrate La proteins (5)
380 WOLIN y CEDERVALL
 THE LA PROTEIN 381

(Figure 1). Other functions that have been mapped to the C-terminal domain of
human La include a nuclear localization signal (62), a nuclear retention signal
(62), a potential dimerization domain (81), and a possible caspase cleavage site
(66).
Metabolic labeling experiments have revealed that La proteins from humans
to yeast are phosphorylated in vivo (10, 82, 83). The human La protein is
phosphorylated at multiple sites, all of which appear to be located within the
C-terminal domain (73, 82, 83). To date, four phosphorylation sites have been
mapped in the human protein. The major site of phosphorylation is a casein
kinase II site at serine 366 (36, 84). In addition, the protein is phosphorylated at
threonine 302, serine 325, and threonine 362 (84). On the basis of sequence
comparison, none of these sites appear conserved beyond vertebrate La proteins.
Consistent with a lack of conservation, the major sites of phosphorylation were
recently mapped in the S. cerevisiae La protein. Two of the three major sites of
phosphorylation map to the N terminus of the protein, and the major C-terminal
site does not correspond to a casein kinase site (59).

The Family of La Motif–Containing Proteins

In addition to the authentic La protein, all sequenced eukaryotic genomes contain
one or more La motif– containing proteins. Outside the La motif, these proteins
are unrelated in amino acid sequence to bona fide La proteins (7, 56, 69, 85)
(Figure 3A). For many, although not all of these proteins, the La motif is located
either centrally or toward the C terminus, in contrast to the N-terminal location
that is characteristic of authentic La proteins (56). Sequence analyses reveal that
the La motifs of these proteins are more similar to each other than those of bona
fide La proteins (56) (see Figure 2 and Figure 3B). Interestingly, in some of these
proteins, the La motif abuts an RRM, just as in authentic La proteins (Figure 3A).

4™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™
Figure 2 Alignment of La motifs from authentic La proteins and other La motif–
containing proteins. Authentic La proteins from humans, Drosophila, S. pombe, S.
cerevisiae, and T. brucei are shown (4, 7–12). C. elegans C44E4.4 (AAB54169) has
not been demonstrated to bind nascent RNA polymerase III transcripts but is
homologous to authentic La proteins throughout its length. Euplotes aediculatus p43,
a component of telomerase, resembles authentic La proteins in overall structure (78).
All La motif– containing proteins in the S. cerevisiae, D. melanogaster, and C.
elegans genomes are included in the lineup, along with related proteins in humans. La
motif– containing proteins in the alignment are human KIAA0217 (accession
XP 040265), FLJ11196 (AAH09446), and KIAA0731 (78a), D. melanogaster
CG11505, AAF27644, CG17386 (78b), and larp (69), C. elegans T12F5.5
(AAB96747) and R144.7 (AAK93859), and S. cerevisiae Sro9p (78c) and Slf1p (87).
Black shading indicates identity in the majority of sequences. Boxed residues are
similar in at least half the sequences. Similar residues were defined as D⫽E,
H⫽K⫽R, I⫽L⫽V, F⫽W⫽Y, S⫽T.
382 WOLIN y CEDERVALL
 THE LA PROTEIN 383

Despite the fact that these La motif– containing proteins are largely unrelated
in primary sequence (apart from the La motif) to authentic La proteins, there is
some phylogenetic conservation among members of the group. For example,
Drosophila larp, a La motif– containing protein that is a target of homeotic genes,
has potential homologs in humans, C. elegans, and the rice plant Oryza sativa
(69) (also Figure 3A).
What roles might La motif proteins play in cells? To date, the only functional
studies of these proteins have been carried out in yeast. The two S. cerevisiae La
motif– containing proteins, Sro9p and Slf1p, are highly related in sequence and
may have resulted from a chromosomal duplication (56). Yeast strains lacking
Sro9p, Slf1p, and the La protein Lhp1p grow similarly to strains lacking only
Sro9p (which have a slight growth defect), indicating that the three proteins do
not function in a single essential process (56). Moreover, whereas the authentic
La protein Lhp1p is nucleoplasmic (55, 56), Sro9p is cytoplasmic (56, 85). Both
Sro9p and Slf1p were found to be RNA-binding proteins that associated with
polyribosomes (56). Since mRNA stability and translation are often coupled,
Sro9p and Slf1p could function in either protein synthesis or mRNA stability
(56). Interestingly, overexpression of Sro9p was found to increase mRNA
stability in vivo (70). Consistent with a general role in mRNA stability and/or
translation, Sro9p and Slf1p have been isolated as high copy suppressors of
mutations in a range of biological processes, including the secretory pathway, the
actin cytoskeleton, copper metabolism, pre-mRNA splicing, and RNA polymer-
ase II transcription (56, 70, 85– 87).
As the La motif is within the region of authentic La proteins required for
specific RNA binding (see below), and as Sro9p and Slf1p are RNA-binding
proteins, one possibility is that the La motif– containing proteins constitute a new

4™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™
Figure 3 Relationships between La and La motif– containing proteins. (A) Mem-
bers of the La motif– containing family of proteins are depicted. All La motif–
containing proteins in the S. cerevisiae, D. melanogaster, and C. elegans genomes are
shown, along with related proteins in humans. The T. brucei La protein (11, 12)
and the Euplotes aediculatus p43 (78) are also displayed. The La motif is in black,
and the RRMs are indicated. Boxed areas with vertical stripes indicate a region of
⬃400 amino acids that is 35– 47% identical between Drosophila larp, C. elegans
R144.7, and human KIAA0731 (56, 69). Boxed areas with horizontal stripes indicate
regions of similarity between human FJL11196, D. melanogaster CG17386, and D.
melanogaster AAF27644. (B) A phylogenetic tree of the La motifs is shown. La
motifs were aligned by MegAlign using the CLUSTAL method (DNASTAR,
Madison, Wisc.). The Paupsearch and Paupdisplay programs (Genetics Computer
Group, Madison, Wisc.) produced the phylogenetic tree using a heuristic search
algorithm and maximum parsimony criterion. Bootstrap analysis based on 1000
replicates was used to assess the reliability of the branching topology. Branches
maintained in 60% or more of replicates are indicated by closed circles.
384 WOLIN y CEDERVALL

family of RNA-binding proteins. If so, identification of their target RNAs may

yield insights into the functions of the individual proteins and also help elucidate
the precise contribution that the La motif makes to RNA recognition by the La
protein.

FUNCTIONS OF THE LA PROTEIN

Both genetic and biochemical studies have revealed that a major role of La is to
protect the 3⬘ end of nascent small RNAs from exonuclease digestion (30, 31, 34,
39 – 41, 44, 45). This La protein–mediated stabilization has different conse-
quences for the various RNAs bound. For example, binding by La to the 3⬘ end
of pre-tRNAs influences the order and mechanism by which the 3⬘ trailer is
removed (39) and may also modulate removal of the 5⬘ leader by RNase P (44,
46). For other small RNAs, binding by La can facilitate RNP assembly (30, 40),
contribute to retaining the nascent RNA in the nucleus (47– 49), or stabilize RNA
structure (39). Although less is known about the roles played by the La protein
in processes involving larger RNAs, La has also been proposed to facilitate
translation of certain cellular and viral-encoded mRNAs (50, 88 –95).

La Is Required for the Normal Pathway of Pre-tRNA

Maturation
In the yeasts S. cerevisiae and S. pombe, La is not essential for viability (7, 9, 10),
making it possible to examine RNA profiles in cells lacking the protein. Yeast
cells lacking La show characteristic changes in the pattern of pre-tRNAs, as
revealed by probing Northern blots (10, 39, 46). All tRNAs are synthesized with
5⬘ and 3⬘ extensions that must be removed to generate mature tRNAs. Analyses
of pre-tRNA maturation in S. cerevisiae revealed that both the order and
mechanism by which the 5⬘ and 3⬘ extensions of many pre-tRNAs are removed
is altered in cells and extracts lacking the La protein (39) (Figure 4). In wild-type
cells, the first processing event is removal of the 5⬘ leader sequence by the
endonuclease RNase P. The La-bound 3⬘ trailer is subsequently removed by an
as yet unidentified endonuclease. In cells lacking La, the first processing event is
nibbling of the 3⬘ trailer by exonucleases (39). Exonuclease digestion halts near
a stem formed in most pre-tRNAs by base-pairing between the purine-rich 5⬘
trailers and U-rich 3⬘ trailers (96, 97). Following RNase P cleavage of the 5⬘
leader, further exonuclease digestion generates the mature 3⬘ end (39) (Figure 4).
Experiments in Xenopus extracts that were immunodepleted of La (41) and
human fractionated cell extracts lacking La (44) confirmed that pre-tRNAs
undergo exonucleolytic nibbling of the 3⬘ end in the absence of La protein. Thus,
in organisms from yeast to humans, binding by La to the ends of pre-tRNAs
protects the 3⬘ trailer from exonucleolytic digestion. Furthermore, at least in
yeast cells, binding by La favors removal of the 3⬘ trailer by an endonucleolytic
cleavage, thus influencing the pathway of pre-tRNA maturation (10, 39).
 THE LA PROTEIN 385

Figure 4 Model for pre-tRNA maturation in wild-type cells and cells lacking the La
protein. In wild-type cells, the nascent pre-tRNA is bound by the La protein. Following
removal of the 5⬘ leader sequence by RNase P, an as yet unidentified endonuclease
removes the La-bound 3⬘ trailer. In cells lacking the La protein, exonucleases digest the
3⬘ trailer up to a helix formed by base-pairing the 5⬘ and 3⬘ extensions. Following RNase
P cleavage, additional exonuclease digestion generates a fully trimmed 3⬘ end. For those
pre-tRNAs that contain introns, removal of the intervening sequence is required to
generate mature tRNA. (Adapted from Reference 39.)

Although the La protein is not required for formation of functional mature

tRNA in otherwise wild-type yeast (10, 39), La becomes required in the presence
of certain mutations in essential tRNA genes or components of the tRNA
biogenesis pathway. For example, mutations in the essential tRNA genes encod-
Ser Arg Thr
ing tRNACGA , tRNACCG , and tRNACGU cause the S. cerevisiae La protein to
become required for viability (39; SD Kim & SL Wolin, unpublished data). La
also becomes essential in S. cerevisiae cells lacking the tRNA(1-methylad-
enosine) methyltransferase, which methylates the adenosine at position 58 in the
T⌿C loop of many eukaryotic tRNAs (45, 98, 99). Interestingly, the La protein
becomes required for different aspects of pre-tRNA biogenesis in each mutant
strain. In cells lacking the tRNA(1-methyladenosine) methyltransferase, La is
required to stabilize pre-tRNAMet
i from degradation (45), which suggests that La
386 WOLIN y CEDERVALL

functions redundantly with the 1-methyladenosine modification to stabilize

pre-tRNAMeti . However, a mutation that disrupts the anticodon stem of pre-
Ser
tRNACGA causes cells to require La for maturation of the mutant pre-tRNA (39).
In the absence of La the mutant pre-tRNA is stable, but is not efficiently
processed to mature tRNA (39). In contrast, a mutation that disrupts the
Arg
anticodon stem of tRNACCG causes La to be required for production of amino-
Arg
acylated tRNA. In this case, the mutant pre-tRNACCG is processed to mature
tRNA, but is not efficiently charged in the absence of La protein (SD Kim & SL
Wolin, unpublished data). One explanation for the different outcomes is that
binding by La to nascent pre-tRNAs favors formation of the correctly folded
RNAs, rather than misfolded alternative structures. In this scenario, the different
outcomes (degradation, failure to be processed, and failure to undergo charging
by synthetases) would be consequences of the particular structure adopted by the
misfolded pre-tRNA. Consistent with the hypothesis that binding by the La
protein may stabilize pre-tRNA structure, introduction of a second mutation that
Ser
restores base-pairing in the anticodon stem of pre-tRNACGA eliminates the
requirement for La in the mutant strain (39).
Recently, it was proposed that phosphorylation of the human La protein
modulates removal of the 5⬘ leader by RNase P (44, 46). In these experiments,
addition of recombinant La protein to pre-tRNA processing reactions was found
to inhibit cleavage by RNase P. The inhibition was not detected when a
phosphorylated form of human La (containing phosphoserine at residue 366) was
assayed (44). Experiments in which the human La protein was expressed in S.
pombe revealed that the human protein blocked maturation of a pre-tRNA
reporter. Since the wild-type human protein was less inhibitory to maturation
than a mutant protein that could not be phosphorylated at S366, phosphorylation
of the human protein was proposed to modulate RNase P cleavage (46).
However, experiments in yeast cells and extracts have failed to reveal a role for
either the S. cerevisiae or S. pombe La protein in modulating RNase P cleavage
(10, 39, 46, 59). Most recently, all major sites of phosphorylation in the S.
cerevisiae protein were identified and mutated. Despite the lack of La protein
phosphorylation, tRNA maturation was unaffected, indicating that phosphoryla-
tion of La is not required for tRNA maturation in S. cerevisiae (59). One possible
explanation for the conflicting results is that the human protein, which is larger
than the yeast proteins, has acquired additional function(s).

La Stabilizes Other Nascent RNAs from Exonuclease

Digestion
Binding by the La protein also protects other newly synthesized small RNAs
from 3⬘ exonucleases. This has been most clearly demonstrated in yeast, as cells
lacking the La protein show changes in the levels and sizes of certain small RNA
precursors. In S. cerevisiae, binding by La stabilizes specific precursors to both
the spliceosomal U1, U2, U4, U5 RNAs and the small nucleolar RNA (snoRNA)
U3(30, 31) (Figure 5). These RNAs, which are transcribed by RNA polymerase
 THE LA PROTEIN 387

Figure 5 Stabilization of newly synthesized small RNAs by the La protein. La

binds and stabilizes newly transcribed U6 snRNA (20, 40). Subsequent binding of the
polyuridylate tract by the Lsm proteins may displace the bound La protein. In yeast,
the La protein also stabilizes 3⬘ extended precursors to the spliceosomal snRNAs and
the small nucleolar RNA U3 (30, 31). For U4 RNA, the precursor stabilized by La
may be the preferred substrate for Sm protein binding (30). Although the Sm proteins
are shown as a ring in the figure, the individual proteins may assemble on the RNA
in a stepwise fashion (105). The U4 and U6 RNAs subsequently base-pair to form the
U4/U6 snRNP.

II, differ from RNA polymerase III transcripts in that La does not bind the
primary transcript. Instead, the La-bound U snRNA precursors are generated by
RNase III cleavage of the primary transcript, followed by exonucleolytic diges-
tion to a run of uridylates (30, 31, 100 –103). Binding by La stabilizes these
precursors, as the nascent RNAs are undetectable or shortened in strains lacking
the La protein (30, 31). Although La is not required for the biogenesis of either
the spliceosomal snRNAs or the U3 snoRNAs in wild-type cells, binding by La
388 WOLIN y CEDERVALL

may slow maturation of these RNAs, possibly as part of a quality-control

function (31).
While La is not essential for small RNA biogenesis in yeast, certain mutations
in the core proteins of small nuclear ribonucleoproteins (snRNPs) cause cells to
require the La protein. Specifically, mutations in members of the Sm and Sm-like
family of proteins cause S. cerevisiae to require La (30, 40, 104). Members of this
family of proteins share a conserved domain known as the Sm motif. The seven
Sm proteins form a heptameric ring that binds a uridylate-rich sequence in the
RNA polymerase II–transcribed U1, U2, U4, and U5 snRNAs (105). In yeast,
binding of the Sm proteins to newly synthesized snRNAs facilitates 3⬘ processing
of the RNAs and hypermethylation of the 5⬘ cap structure, and also stabilizes the
RNAs from degradation (103, 106 –108). A mutation in SMD1, which encodes
the Sm D1 protein, causes S. cerevisiae cells to require La at low temperatures
(30). In these cells, La becomes required for assembly of the U4 RNA into its
ribonucleoprotein (RNP) form, the U4/U6 snRNP. Since the pre-U4 RNA, but
not the mature U4 RNA, is bound by Smd1p in vitro, La may function in these
cells by stabilizing the preferred substrate for Sm protein binding (30). Similarly,
seven related proteins, known as Lsm (for “Like Sm”) proteins, form a hep-
tameric ring that binds to the polyuridylate stretch at the 3⬘ end of the U6 snRNA
(109 –112). In the presence of mutations in the Lsm protein complex, the La
protein becomes required to stabilize newly synthesized U6 snRNA (40, 104).
Does binding by La also stabilize nascent U snRNAs in metazoans? In
vertebrate cells, approximately 10% of the U6 snRNA is bound by the La protein
(20, 113). The U6 snRNA bound by La likely represents newly synthesized
RNA, as it contains fewer modified nucleotides than the total population (20).
Moreover, whereas the fraction of U6 RNA bound by La terminates with
UUUOH, the majority of U6 RNAs in vertebrate cells contain 2⬘,3⬘ cyclic
phosphates at their 3⬘ ends (113, 114). Thus, as has been demonstrated in yeast,
binding by La may stabilize newly synthesized U6 snRNA in higher cells. It has
been less clear whether La also binds precursors to the RNA polymerase
II–transcribed U1, U2, U4, and U5 snRNAs in metazoans. A population of U1
snRNAs that are longer than mature U1 RNA are bound by the La protein in
vertebrate cells (115). These longer U1 RNAs were reported to be cytoplasmic
(115). However, this may not be surprising, as the vast majority of La-bound
RNAs, including pre-tRNAs and pre-5S rRNA, leak into the cytoplasm during
similar fractionation procedures (13, 29). Thus, as in yeast, La may bind newly
synthesized pre-U1 RNAs in vertebrate cells. However, precursors to U2, U4,
and U5 snRNAs have not yet been described as associating with the La protein
in metazoan cells.

La May Contribute to Nuclear Retention of Nascent

Transcripts
Because the nascent small RNAs bound by the La protein are nuclear, whereas
the mature RNAs are often cytoplasmic, it has long been proposed that binding
 THE LA PROTEIN 389

by La may retain RNA polymerase III transcripts in the nucleus (29, 47– 49). To
test this hypothesis, Boelens et al. (47) examined whether binding by La
contributed to nuclear retention of U6 snRNA. Because La binds the UUUOH at
the 3⬘ end of nascent RNA polymerase III transcripts (see below), the authors
constructed a mutant Xenopus laevis U6 RNA lacking the terminal uridylates.
Interestingly, when the mutant RNA was injected into oocytes, a significant
fraction of the normally nuclear U6 RNA was found in the cytoplasm, which
suggests that La contributes to nuclear retention. However, as only about 10% of
the U6 snRNA in most cell types is associated with La (20, 40, 113), the authors
noted that it was unlikely that La binding caused retention of the majority of U6
snRNAs. Instead, there might be retention factors, distinct from the La protein,
that also recognize the 3⬘ end of U6 snRNA (47). This interpretation turned out
to be remarkably prescient, as it was subsequently discovered that the Lsm
protein complex binds the 3⬘ end of mature U6 snRNA (109, 112). Thus,
truncation of the 3⬘ end of U6 snRNA may have interfered with stable binding
of the Lsm protein complex. Whether binding by Lsm proteins contributes to
nuclear retention of U6 snRNA is not known.
The strongest case for a role for La in nuclear retention comes from studies of
the human Y1 RNA. Although the function of this RNA polymerase III–
transcribed small RNA is unknown, most of the RNA is found in the cytoplasm
bound to the 60-kilodalton Ro protein (14, 60, 61, 116). In addition, a fraction of
the hY1 RNA is bound by the La protein in the nucleus (61). When hY1 RNA
transcribed in vitro is injected into Xenopus oocytes, the RNA is slowly exported
to the cytoplasm, requiring more than eight hours for complete export (48, 117).
However, a mutant hY1 RNA lacking the terminal uridylates was completely
cytoplasmic within two hours (48). Although this experiment does not rule out
the possibility that mutation of the terminal uridylates interfered with the binding
of other nuclear retention factors, the results suggest that binding by La contrib-
utes to nuclear retention of hY1 RNA (48). Support for this idea was provided by
Grimm et al. (49), who used an in vivo selection technique to identify RNA
sequences that were retained in the nucleus. One of the selected RNAs, called
NL-15, was bound by the La protein in oocyte nuclei. Injection of high levels of
NL-15 was found to increase the export of hY1 RNA, perhaps because the excess
NL-15 RNA competed with hY1 RNA for binding to La (49).
If binding by La can retain nascent RNA polymerase III transcripts in the
nucleus, could La binding also retain mature RNPs in this compartment?
Although the overwhelming majority of La-associated small RNAs are precur-
sors, several mature RNPs are almost entirely associated with a La protein. In
ciliates, the RNA component of telomerase is transcribed by RNA polymerase
III. In contrast to most polymerase III–transcribed RNAs, ciliate telomerase
RNAs retain their terminal uridylates in mature, active telomerase. Purification of
the telomerase RNP from the ciliate Euplotes aediculatus revealed that one
protein component of this RNP, p43, is a La motif– containing protein that
resembles a bona fide La protein in its overall structure (78). Nearly all of the E.
390 WOLIN y CEDERVALL

aediculatus telomerase activity was associated with p43, indicating that this
La-like protein is a component of mature telomerase. As p43 has not been
demonstrated to bind nascent RNA polymerase III transcripts in vivo, it is not yet
clear whether it is an authentic La protein or has a more specialized function.
Nonetheless, because of the structural similarities to La proteins, it has been
proposed that binding by p43 retains telomerase in the nucleus (78). Other mature
RNAs that are largely bound by the La protein include the Epstein-Barr
virus-encoded EBER RNAs and herpesvirus papio HVP RNAs (118). Similar to
the ciliate telomerase, binding by the La protein may contribute to nuclear
retention of these RNAs in mammalian cells.
Although there is good evidence that binding by La contributes to nuclear
retention of hY1 and NL-15 RNAs in Xenopus oocytes, it is not yet clear whether
this is a general role of the La protein. As the La protein is dispensable for growth
in both S. cerevisiae and S. pombe (7, 9, 10), La cannot be required for the
nuclear retention of any essential RNA in either yeast. Although a homolog of
hY1 RNA has not been described in yeast, pre-tRNAs are localized to the S.
cerevisiae nucleolus and nucleus (119 –121). Thus, binding by La could poten-
tially contribute to either nuclear or nucleolar localization of these nascent RNAs.
Leu
However, experiments in which two S. cerevisiae pre-tRNAs (pre-tRNACAA and
Ile
tRNAUAU ) were detected using in situ hybridization revealed that their localiza-
tion was unaffected in cells lacking La (H Grosshans, personal communication).
One possibility for the conflicting results is that the vertebrate La protein has
evolved an additional function of retaining certain nascent transcripts, such as
hY1 RNA, in the nucleus. Alternatively, yeast may possess redundant mecha-
nisms for retaining pre-tRNAs in the nucleus. It should also be noted that
Xenopus oocytes are unusual cells, in that large quantities of proteins and RNAs
are stored in preparation for early development. Thus, results obtained in oocytes
may differ from those obtained in somatic cells.

Is La a Transcription Factor for RNA Polymerase III?

One of the earliest proposed roles for the La protein was as a transcription
termination and recycling factor for RNA polymerase III. In these experiments,
human HeLa cell extracts that were immunodepleted of La protein were found to
be drastically reduced in their ability to transcribe polymerase III genes (32). As
the residual transcripts that were synthesized were slightly shorter at the 3⬘ end,
a model was proposed in which La was required for proper termination and
release of RNA polymerase III transcripts (32, 33). In subsequent experiments
using immobilized transcription complexes, it was reported that addition of
purified human La increased transcript release and was required for reinitiation
by RNA polymerase III (35, 38, 122, 123). Moreover, since the phosphorylated
form of La was inactive in stimulating transcription, phosphorylation of La was
proposed to regulate polymerase recycling (36). However, other groups reported
that RNA polymerase III purified from vertebrates was capable of accurate
 THE LA PROTEIN 391

termination and transcript release in the absence of other proteins (124 –126),
making a role for La in transcription controversial.
In recent years, several groups have specifically investigated the role of the La
protein in RNA polymerase III transcription. S. cerevisiae extracts that were
genetically depleted of La were found to be similar to wild-type extracts in their
ability to transcribe tRNA and U6 RNA genes (39, 40). Similarly, Xenopus cell
extracts that were immunodepleted of La protein were comparable to wild-type
extracts for transcribing tRNA genes (41). However, in both the yeast and
Xenopus experiments, the transcripts made in the La-depleted extracts were
shortened at the 3⬘ end by exonucleases, consistent with a role for La in
protecting the 3⬘ ends of nascent transcripts (39 – 41). Most recently, Weser et al.
(42) examined the role of the human La protein in RNA polymerase III
transcription. These authors found that a reconstituted human transcription
system that was immunodepleted of the La protein was fully active in transcrip-
tion (42). Consistent with this result, transcription of human U6 RNA from
partially purified components was found to be efficient in the absence of
detectable La protein (43).
Given that transcription in yeast and human cell extracts is efficient in the
absence of detectable La protein (39 – 43), it seems clear that the La protein is not
required for RNA polymerase III transcription in vitro. Moreover, since the La
protein is dispensable for growth in both S. cerevisiae and S. pombe (7, 9, 10),
La is not required for the transcription of any essential RNA polymerase III RNA
in these yeasts. However, it remains possible that La interfaces with the
transcription machinery to modulate transcription in vivo.

Possible Roles for La in mRNA Translation

A number of studies have implicated the La protein in translation of specific viral
and cellular mRNAs. In the first of these studies, La was identified as the major
protein that could be cross-linked with ultraviolet light to a fragment of the
poliovirus 5⬘ untranslated region (50). Poliovirus, like other picornaviruses, is
translated by cap-independent internal ribosome binding (127). Poliovirus RNA
is poorly translated in rabbit reticulocyte lysates, in that initiation of translation
occurs at aberrant sites in the RNA. Addition of high amounts of La protein
(⬃10-fold greater than that present in a human HeLa cell extract) suppressed
initiation at these aberrant sites, suggesting that La could play a role in
facilitating proper initiation (50). Consistent with a role in translation, the
normally nuclear La protein redistributes to the cytoplasm in poliovirus-infected
cells (50), possibly as a result of cleavage by a poliovirus-encoded protease (63).
Interestingly, transcription of the human La gene results in two transcripts that
differ only in their 5⬘ noncoding regions (128). One of these transcripts contains
an internal ribosome entry site, suggesting that La itself may continue to be
translated under conditions where cap-dependent translation is impaired, such as
poliovirus infection (129).
392 WOLIN y CEDERVALL

La has also been implicated in internal ribosome binding to hepatitis C virus

(HCV) RNA (91, 93). In these experiments, La protein was cross-linked with
ultraviolet light to an RNA fragment containing the 5⬘ noncoding region of HCV.
Interestingly, studies in which pieces of the HCV 5⬘ noncoding region were used
as competitors for cross-linking revealed that only RNAs containing the initiator
AUG codon competed for binding to the La protein. Moreover, addition of La to
a reticulocyte lysate stimulated translation of a reporter protein containing the
HCV 5⬘ noncoding region (91). To determine whether La was required for
translation of HCV, an RNA sequence that was selected for high-affinity binding
to the human La protein was used to sequester La protein. Addition of this RNA
to a rabbit reticulocyte lysate inhibited translation of a reporter gene linked to the
HCV internal ribosome entry site (93). Furthermore, transfection of this RNA
into human cells decreased translation of the same reporter gene (93).
Although human immunodeficiency virus type 1 (HIV-1) does not use internal
ribosome binding for translation, La has also been proposed to facilitate trans-
lation of the virus-encoded mRNAs (89). All HIV-1 mRNAs contain a stem-loop
structure, known as the TAR element, at their 5⬘ ends. This TAR sequence
inhibits translation in vitro, in part because of its stable secondary structure (130).
The La protein binds the TAR RNA, both in vitro and in HIV-1 infected cells
(88). Similar to poliovirus and HCV RNAs, addition of high amounts of La
protein stimulated translation of a reporter gene linked to the TAR element (89).
Although addition of La protein to reticulocyte lysates enhances translation of
these three viral RNAs, the significance of some of these observations has been
questioned (131, 132). One issue is that the quantities of La protein required to
stimulate translation or suppress aberrant initiation are quite large (⬃2 ␮M for
poliovirus), which suggests that high amounts of La protein could be substituting
for a physiologically relevant RNA-binding protein (131). Consistent with this
idea, addition of several other RNA-binding proteins [heterogeneous nuclear
(hn)RNP I/PTB, hnRNP A1, and p50] also suppressed initiation of poliovirus
translation at aberrant sites (133). A second issue is that for both poliovirus and
HCV, La has not been demonstrated to bind the endogenous RNAs in cells. A
final concern is that immunodepletion experiments to test the requirement for La
in these systems have not succeeded. In one such experiment, immunodepletion
of La from a HeLa cell lysate eliminated translation. However, activity could not
be restored by addition of purified La protein (134), making it unclear whether
La or an associated protein was required. Nonetheless, the result that an RNA that
binds La with high affinity inhibits translation of RNAs containing the HCV
noncoding region (93) supports the idea that La may contribute to start site
selection, at least for this virus.
Despite the fact that La is largely nuclear in uninfected cells, the vertebrate
protein has also been implicated in the translation of several cellular mRNAs. La
was cross-linked with ultraviolet light to the 5⬘ untranslated region of a cellular
mRNA that is translated by internal ribosome binding, the X-linked inhibitor of
apoptosis protein (XIAP) (95). This mRNA could be detected in anti-La immu-
 THE LA PROTEIN 393

noprecipitates from cultured cells, although detection required overexpression of

the XIAP mRNA (95). Experiments performed in reticulocyte lysates revealed
that a C-terminal La fragment that acts as a dominant negative in inhibiting
poliovirus translation in vitro (81) also inhibited translation of a reporter gene
containing the XIAP 5⬘ noncoding region (95). During apoptosis, the C terminus
of La is cleaved by a caspase-dependent mechanism, removing the nuclear
localization signal (65, 66). Thus, similar to the situation in virus-infected cells,
the normally nuclear La protein may become cytoplasmic during apoptosis (65,
66), consistent with a role in influencing translation of XIAP mRNA.
The La protein has also been proposed to regulate the translation of mRNAs
containing 5⬘ terminal oligopyrimidine sequences (TOP mRNAs). This interest-
ing class of mRNAs, which encode ribosomal proteins and certain other com-
ponents of the translational apparatus, are regulated at the translational level in
a growth-dependent manner (135). In growth-arrested cells, translation of these
mRNAs is repressed, as evidenced by decreased polyribosome association (135).
La was implicated in the translational regulation of these mRNAs by the finding
that the Xenopus protein could be cross-linked to a short RNA (12 nucleotides)
consisting solely of the oligopyrimidine sequence (92, 136). Although La has not
been demonstrated to associate with TOP mRNAs in vivo, cell lines that
overexpress Xenopus La show a small increase in the fraction of ribosomal
protein mRNAs found on polyribosomes during growth arrest (94). However, in
similar experiments, others failed to detect any effect of La overexpression on
TOP mRNA translation (135). In addition, since the TOP mRNAs are among the
most abundant mRNAs in vertebrate cells (135), it is unclear how the small
fraction of La in the cytoplasm would modulate translation of this large class of
abundant mRNAs.
Could the La protein play a general role in translation? The human La protein
has been reported to sediment with 40S ribosomal subunits and to bind 18S
rRNA (137). In addition, site-specific ultraviolet cross-linking experiments in
human cell extracts have revealed that La can be cross-linked to a start-site AUG
codon, but only when the AUG is embedded in a favorable Kozak consensus
sequence (138). However, experiments in which La was added to a rabbit
reticulocyte lysate failed to reveal a role for the protein in start-site selection
(138). Complicating the question of whether La plays a role in translation is the
finding that addition of high concentrations (approximately 3.2 ␮M) of La or
several other general RNA-binding proteins [hnRNP A1, hnRNP I/PTB, and the
messenger ribonuleoprotein (mRNP) p50] to reticulocyte lysates significantly
decreases the translation of uncapped, but not capped, mRNAs (133). Since La,
hnRNP A1, and hnRNP I/PTB are all predominantly nuclear in uninfected
mammalian cells, the authors suggested that a cytoplasmic general RNA-binding
protein, such as p50, may normally fulfill this function in vivo (133). Thus,
although evidence has begun to accumulate that binding by the La protein
facilitates translation of specific viral and cellular RNAs, at present there is little
functional evidence for a general role for La in translation initiation.
394 WOLIN y CEDERVALL

Other Potential Roles for the La Protein

In addition to the proposed roles for the La protein in translation, La has been
implicated in a number of other processes. For some of these processes, La was
identified as a protein in cell extracts that could be cross-linked or otherwise
bound to a target RNA. For example, the human La protein was identified as the
protein that bound and stabilized a fragment of histone mRNA (that terminated
in uridylates) from exonuclease digestion (139). A role for the La protein in
binding and stabilizing UUUOH-containing mRNA fragments from exonucleases
would be similar to the role of the La protein in stabilizing UUUOH-containing
snRNA precursors in yeast (30, 31). However, La has not yet been demonstrated
to bind histone mRNA degradation intermediates in vivo. Similarly, the La
protein was identified in several laboratories as binding to short RNA sequences
derived from viruses (140 –143), and proposed to function in various aspects of
the viral life cycles, including replication (140) and stabilization of viral RNA
(142, 143). However, a functional role for La in these processes has not yet been
demonstrated.

RNA BINDING BY THE LA PROTEIN

RNA Sequences Recognized by La

The vast majority of small RNAs bound by the La protein in cells terminate in
the sequence UUUOH. Although most of these RNAs are nascent RNA polymer-
ase III transcripts (14, 15, 20), La also binds to RNAs made by other polymerases
that end in UUUOH (24, 30, 31). A number of studies have established the
importance of the terminal uridylates for high-affinity binding by the La protein.
First, for both human 5S rRNA and adenovirus VA RNAI, the form of the RNA
bound by the La protein differs from the unbound form only in having additional
uridylates at the 3⬘ end (15, 144). Second, ribonuclease protection experiments
performed on immunoprecipitated rat 4.5S RNA (28) and Xenopus pre-5S rRNA
(145) revealed that bound La protein protects the 3⬘ ends of these RNAs from
digestion. Moreover, the human La protein can be cross-linked with ultraviolet
light to the 3⬘ end of adenovirus VA RNAI (144). Lastly, a systematic study using
purified human protein and model RNA substrates revealed that La preferentially
binds RNAs terminating in three uridylate residues (29). Conversion of the
terminal hydroxyl to a phosphate reduced the binding of the human protein 10-
to 20-fold, demonstrating the importance of the 3⬘ hydroxyl for high-affinity
binding (29). Similar experiments, in which the 3⬘ hydroxyl of U6 snRNA was
converted to a 3⬘ phosphate, demonstrated that La proteins from Xenopus laevis,
Drosophila melanogaster, and Saccharomyces cerevisiae all preferentially bind
RNAs containing a 3⬘ hydroxyl (7, 59, 113).
Although the 3⬘ uridylates are a major determinant for La protein binding, the
protein must also recognize other features of RNA structure. This was first noted
 THE LA PROTEIN 395

by Stefano (29), who observed that a model pre-tRNA substrate (a mature tRNA
containing additional uridylates ligated to the 3⬘ end) was bound less stably by
La than a bona fide pre-tRNA. Also, some of the RNAs that associate with the
La protein in cells do not end in uridylates (25, 26, 88). Although the other
features of La-bound RNAs that contribute to recognition by La have not been
systematically studied, the vertebrate La protein binds certain long stem struc-
tures lacking uridylates (49, 88). For one case, that of the human immunodefi-
ciency virus (HIV) TAR RNA, mutations that disrupted the stem structure
eliminated binding (88). Thus, one possibility is that the extended acceptor stems
present in all pre-tRNAs, due to base-pairing between the purine-rich 5⬘ leaders
and pyrimidine-rich 3⬘ trailers (97), contribute to stable binding by La. The
human La protein also binds a pre-tRNA containing a 5⬘ triphosphate two- to
threefold more efficiently than the identical dephosphorylated RNA (44). Thus,
La may also recognize the 5⬘ triphosphate that is at the end of all newly
synthesized RNA polymerase III transcripts. However, as this difference in
binding became apparent upon removal of the 3⬘ uridylates from the pre-tRNA,
the contribution of the 5⬘ triphosphate to overall recognition by the La protein
binding may be small compared to the terminal uridylates (44). It has also been
proposed that the La protein may recognize internal oligouridylates (146);
however, this has not been demonstrated experimentally.

Features of La That Contribute to RNA Recognition

Although it is not yet known which surfaces of the La protein are in direct contact
with RNA, deletion analyses have defined the regions of the human La protein
that are required for RNA binding. These experiments revealed that N-terminal
fragments of the human protein, consisting of the La motif, the first RRM, and
⬃17 adjacent amino acids, bind several RNA substrates with an affinity com-
parable to that of the full-length protein (74, 76, 123, 146). Competition
experiments revealed that this La motif–RRM1 fragment retains the ability to
discriminate between a polyuridine substrate and a nonspecific competitor (76).
Thus, the C terminus, including the atypical second RRM found in vertebrate La
proteins, is not required for either RNA binding affinity or specificity.
Both the La motif and the first RRM appear critical for RNA binding by the
human protein. Although the isolated La motif does not bind RNA, even small
deletions within the motif drastically decrease RNA binding affinity (74, 88,
123). However, at high protein concentrations, forms of the human La protein
lacking the La motif still bind RNA (123), which suggests that at least some
general RNA binding ability is conferred by the RRM alone. Thus, the La motif
may increase the affinity of the RRM for RNA (123). However, since competi-
tion experiments to examine RNA binding specificity were not performed with
these truncation mutants, it is not known whether the isolated RRM possesses the
specificity for UUUOH that is characteristic of the intact protein. Thus a second,
but not mutually exclusive possibility is that the La motif confers binding
specificity for UUUOH (74, 75).
396 WOLIN y CEDERVALL

Although the C terminus of the human La protein is dispensable for specific

binding of UUUOH-containing substrates, sequences within this domain may
contribute to RNA recognition. Addition of either the C-terminal RRM or the
entire C-terminal domain to the N-terminal La motif–RRM1 fragment decreases
binding of the human La protein to polyuridine (76), which suggests that the
second RRM in the C-terminus modulates RNA binding (76). The C terminus of
vertebrate La proteins also contains a potential Walker A nucleoside triphos-
phate– binding motif (5); however, ATP and nonhydrolyzable ATP analogs were
found to have no effect on the affinity of the human protein for polyuridine (76).
Instead, this sequence has been proposed to contact the triphosphate found at the
5⬘ end of all nascent RNA polymerase III transcripts (44, 46, 75). In these
experiments, a panel of truncated La proteins was examined for the ability to
protect a pre-tRNA 5⬘ leader sequence from cleavage by RNase P. As La proteins
lacking a C-terminal basic region that includes the potential Walker A motif were
inactive in this assay, it was proposed that the Walker A motif contacts the 5⬘
triphosphate (44). A direct interaction between the 5⬘ triphosphate and the
Walker A motif has not yet been demonstrated. Interestingly, although La
proteins from yeast to humans bind pre-tRNA, the potential Walker A motif
sequence GXXXXXGKX is found only in vertebrate La proteins. Since other La
proteins have not been reported to protect pre-tRNAs from RNase P cleavage,
one possibility is that only the vertebrate proteins contact the 5⬘ triphosphate.
Alternatively, since other amino acids within the basic region are conserved in
invertebrates (7, 10), these residues, rather than the Walker motif, could poten-
tially contact the 5⬘ triphosphate.
Since the C terminus of the human protein contains multiple phosphorylation
sites (36, 73, 82– 84), several groups have examined whether phosphorylation
modulates RNA binding. In one study, phosphorylation of the human protein had
little effect on RNA binding affinity (36), while another study reported a twofold
decrease in binding affinity upon phosphorylation with casein kinase (76). Thus,
phosphorylation may have at best a small effect on the binding affinity of the
human La protein. Consistent with this, phosphorylated and unphosphorylated
forms of the S. cerevisiae La protein were found to be very similar in RNA
binding affinity (59). However, the phosphorylated yeast protein exhibited
slightly higher specificity than the unphosphorylated form in discriminating
between RNAs ending in UUUOH and UUp, suggesting that phosphorylation may
make a small contribution to RNA binding specificity (59).
How does the La protein contact RNAs lacking a 3⬘-UUUOH terminus? For
some of these RNAs, such as the HIV TAR RNA and the poliovirus 5⬘
untranslated region, the portions of the human La protein that are required for
binding are similar to those required for binding UUUOH-containing substrates
(88, 90). For both the HIV and poliovirus RNAs, only the N-terminal half of the
La protein, containing the La motif and the first RRM, is required for La binding
(88, 90). However, at least for the TAR RNA, the binding affinity of La is
reduced nearly threefold compared to a UUUOH-containing RNA, the hY4 RNA
 THE LA PROTEIN 397

(88). Competition experiments revealed that whereas La binding to hY4

RNA was competed most effectively by excess poly(U), La binding to the TAR
RNA was most sensitive to excess poly(G) (88). Thus, while the regions of La
that are required to bind hY4 RNA and the TAR RNA are similar, there are likely
to be some differences as to how La contacts these RNAs. Interestingly, for at
least one non-UUUOH-containing RNA, the internal ribosome entry site of the
hepatitis C virus, RNA binding requires only the C-terminal half of the protein
(93). Thus, the mechanism by which La recognizes this RNA may involve the
atypical RRM found in the C terminus of vertebrate La proteins.

Is Oligomerization Required for La Function?

Some evidence suggests that La forms higher-order complexes that may be
important for function. Experiments in which radiolabeled human La protein was
used to probe Western blots revealed that La could interact with itself in vitro
(81). Moreover, using size exclusion chromatography, the purified protein eluted
at a measured molecular weight consistent with dimer formation (81). Experi-
ments in which truncated forms of the human protein were used in the Far-
Western assay revealed that sequences in the C-terminal domain (between amino
acids 293 and 348) were required for homodimer formation (81). These
sequences include part of the atypical RRM found in the C terminus of vertebrate
La proteins, as well as the adjacent basic region and potential Walker A motif.
Interestingly, these same sequences are required for the activity of La in
enhancing in vitro translation of mRNAs containing the internal ribosome entry
sites of poliovirus and hepatitis C virus (81, 93) and mRNAs containing the HIV
TAR element (81). Addition of a fragment of the La protein containing the
dimerization element to a translation reaction reduced the stimulatory effect of
the full-length protein, which suggests that dimerization is important for the
activity of La in translation (81).
Additional evidence for oligomerization comes from studies of RNA binding.
When high concentrations of La are used in electrophoretic mobility shift assays,
additional complexes, of lower mobility than the primary complexes, are
detected on native gels (44, 59, 76, 123). Similar multimeric complexes have
been detected with several other RRM-containing RNA-binding proteins, and
can be due either to dimer formation or to the binding of additional protein
molecules to other, possibly less specific, sites on the RNA target (147, 148).
Although a systematic study of the La protein– containing multimeric complexes
has not been reported, both protein oligomerization and binding to less specific
RNA targets may be involved. First, an additional complex was detected when
the binding of the human protein to a 9-meric uridine oligonucleotide RNA
substrate was studied (76). Since it is unlikely that two copies of La could
simultaneously bind the 9-mer, the slower migrating complex was likely due to
oligomerization. This additional complex was not detected when a truncated La
protein containing only the N-terminal La motif and the first RRM was used in
the binding experiment (76), but appeared when a longer version of La,
398 WOLIN y CEDERVALL

containing the extra RRM in the C terminus, was assayed (76). Thus, the
C-terminal RRM may function in protein-protein interactions. This would not be
unprecedented, as several RRMs have been demonstrated to function in protein
oligomerization (149, 150).
When increasing concentrations of La protein are mixed with larger RNA
substrates, such as hY4 RNA or pre-tRNA, a series of slower-mobility com-
plexes is also detected on native gels (44, 59, 123; KS Long & SL Wolin,
unpublished data). Competition experiments using unlabeled RNA reveal that
significantly less competitor RNA is needed to disrupt these complexes than the
primary complexes, indicating that the binding of La in the slower-mobility
complexes is of lower specificity (59). Experiments using truncated forms of the
human protein revealed that the slower-mobility complexes do not form on hY4
RNA when regions C-terminal to the second RRM were deleted (123). As this
portion of the C terminus includes the basic region, one possibility is that with
increasing amounts of protein, a second La protein molecule interacts via the
basic region with additional, less specific sites on the hY4 RNA. Whether these
lower-specificity interactions are important for La protein function is not known.
However, as some activities attributed to the La protein require the addition of
very large amounts of protein, the contribution of these lower-specificity inter-
actions may warrant investigation.

PERSPECTIVES AND FUTURE DIRECTIONS

In the 20 years since the La protein was first demonstrated to bind nascent small
RNAs, much has been learned about its RNA targets and functions in cells. A
combination of genetic and biochemical studies has revealed that La protein
binding stabilizes nascent RNAs, thus facilitating pre-tRNA maturation and
assembly of small RNAs into functional RNPs. Binding by the La protein also
retains certain nascent small RNAs in the nucleus. Although still controversial,
evidence has also begun to accumulate that binding by the La protein to specific
mRNAs may facilitate initiation of translation.
The challenge for the future is to understand the molecular mechanisms by
which La protein binding to its RNA targets facilitates their biogenesis and
cellular functions. For example, since the La protein is the first protein that binds
to many small RNAs, binding by La could potentially facilitate RNA folding or
resolve misfolded intermediates. One obvious goal is the generation of high-
resolution structures of the La protein bound to its various RNA targets.
Biophysical techniques that allow the monitoring of RNA conformational
changes may also be useful in determining whether binding by the La protein
affects RNA folding. From a cell biological perspective, it is interesting to note
that many, if not all, nascent small RNAs transit through the nucleolus as part of
their normal biogenesis (151). Binding by the La protein could assist either in
nucleolar targeting or in the RNA modification and RNP assembly events that
 THE LA PROTEIN 399

occur within this compartment (152). Moreover, it is unlikely that all the RNAs
bound by the La protein have been uncovered. The application of new genome-
wide methods such as microarray analyses (153, 154) to identify the entire
spectrum of RNAs bound by the La protein seems certain to reveal new insights
into the functions of this highly abundant and ubiquitous RNA-binding protein.

ACKNOWLEDGMENTS
We thank Anne Marie Quinn for assistance with the phylogenetic analyses. We
are grateful to Helge Grosshans, Ger Pruijn, Yousif Shamoo, and Elisabetta Ullu
for sharing unpublished data. We also thank Stefan Aigner, Carl Hashimoto, and
Elisabetta Ullu for critical reading of the manuscript. Work in the Wolin
laboratory on the La protein is supported by NIH grant R01-GM48410. TC was
supported by STINT, the Swedish Foundation for International Cooperation in
Research and Higher Education. SLW is an Associate Investigator of the Howard
Hughes Medical Institute.

The Annual Review of Biochemistry is online at http://biochem.annualreviews.org

LITERATURE CITED
1. Mattioli M, Reichlin M. 1974. Arthritis
Rheum. 17:421–29
2. Alspaugh MA, Tan EM. 1975. J. Clin.
Invest. 55:1067–73
3. Chambers JC, Keene JD. 1985. Proc.
Natl. Acad. Sci. USA 82:2115–19
4. Chambers JC, Kenan D, Martin BJ,
Keene JD. 1988. J. Biol. Chem. 263:
18043–51
5. Topfer F, Gordon T, McCluskey J. 1993.
J. Immunol. 150:3091–100
6. Scherly D, Stutz F, Lin-Marq N, Clark-
son SG. 1993. J. Mol. Biol. 231:196 –204
7. Yoo CJ, Wolin SL. 1994. Mol. Cell.
Biol. 14:5412–24
8. Bai CY, Li ZH, Tolias PP. 1994. Mol.
Cell. Biol. 14:5123–29
9. Lin-Marq N, Clarkson SG. 1995. J. Mol.
Biol. 245:81– 85
10. Van Horn DJ, Yoo CJ, Xue D, Shi H,
Wolin SL. 1997. RNA 3:1434 – 43
11. Marchetti MA, Tschudi C, Kwon H,
Wolin SL, Ullu E. 2000. J. Cell Sci.
113:899 –906
12. Westermann S, Weber K. 2000. Bio-
chim. Biophys. Acta 1492:483– 87
13. Lerner MR, Boyle JA, Hardin JA, Steitz
JA. 1981. Science 211:400 –2
14. Hendrick JP, Wolin SL, Rinke J, Lerner
MR, Steitz JA. 1981. Mol. Cell. Biol.
1:1138 – 49
15. Rinke J, Steitz JA. 1982. Cell 29:149 –59
16. Hashimoto C, Steitz JA. 1983. J. Biol.
Chem. 258:1379 – 82
17. Chambers JC, Kurilla MG, Keene JD.
1983. J. Biol. Chem. 258:11438 – 41
18. Reddy R, Tan EM, Henning D, Nohga
K, Busch H. 1983. J. Biol. Chem. 258:
1383– 86
19. Shen CK, Maniatis T. 1982. J. Mol.
Appl. Genet. 1:343– 60
20. Rinke J, Steitz JA. 1985. Nucleic Acids
Res. 13:2617–29
21. Rosa MD, Gottlieb E, Lerner MR, Steitz
JA. 1981. Mol. Cell. Biol. 1:785–96
22. Lerner MR, Andrews NC, Miller G,
Steitz JA. 1981. Proc. Natl. Acad. Sci.
USA 78:805–9
400 WOLIN y CEDERVALL
23. Kurilla MG, Cabradilla CD, Holloway
BP, Keene JD. 1984. J. Virol. 50:773–78
24. Kurilla MG, Keene JD. 1983. Cell
34:837– 45
25. Wilusz J, Kurilla MG, Keene JD. 1983.
Proc. Natl. Acad. Sci. USA 80:5827–31
26. Wilusz J, Keene JD. 1984. Virology 135:
65–73
27. Francoeur AM, Mathews MB. 1982.
Proc. Natl. Acad. Sci. USA 79:6772–76
28. Reddy R, Henning D, Tan E, Busch H.
1983. J. Biol. Chem. 258:8352–56
29. Stefano JE. 1984. Cell 36:145–54
30. Xue D, Rubinson DA, Pannone BK, Yoo
CJ, Wolin SL. 2000. EMBO J. 19:
1650 – 60
31. Kufel J, Allmang C, Chanfreau G, Pet-
falski E, Lafontaine DL, Tollervey D.
2000. Mol. Cell. Biol. 20:5415–24
32. Gottlieb E, Steitz JA. 1989. EMBO J.
8:841–50
33. Gottlieb E, Steitz JA. 1989. EMBO J.
8:851– 61
34. Maraia RJ, Kenan DJ, Keene JD. 1994.
Mol. Cell. Biol. 14:2147–58
35. Maraia RJ. 1996. Proc. Natl. Acad. Sci.
USA 93:3383– 87
36. Fan H, Sakulich AL, Goodier JL, Zhang
X, Qin J, Maraia RJ. 1997. Cell
88:707–15
37. Chu WM, Ballard RE, Schmid CW.
1997. Nucleic Acids Res. 25:2077– 82
38. Goodier JL, Maraia RJ. 1998. J. Biol.
Chem. 273:26110 –16
39. Yoo CJ, Wolin SL. 1997. Cell 89:393–
402
40. Pannone BK, Xue D, Wolin SL. 1998.
EMBO J. 17:7442–53
41. Lin-Marq N, Clarkson SG. 1998. EMBO
J. 17:2033– 41
42. Weser S, Bachmann M, Seifart KH,
Meissner W. 2000. Nucleic Acids Res.
28:3935– 42
43. Chong SS, Hu P, Hernandez N. 2001.
J. Biol. Chem. 276:20727–34
44. Fan H, Goodier JL, Chamberlain JR,
Engelke DR, Maraia RJ. 1998. Mol. Cell.
Biol. 18:3201–11
45. Calvo O, Cuesta R, Anderson J, Gutier-
rez N, Garcia-Barrio MT, et al. 1999.
Mol. Cell. Biol. 19:4167– 81
46. Intine RV, Sakulich AL, Koduru SB,
Huang Y, Pierstorff E, et al. 2000. Mol.
Cell 6:339 – 48
47. Boelens WC, Palacios I, Mattaj IW.
1995. RNA 1:273– 83
48. Simons FHM, Rutjes SA, van Venrooij
WJ, Pruijn GJM. 1996. RNA 2:264 –73
49. Grimm C, Lund E, Dahlberg JE. 1997.
EMBO J. 16:793– 806
50. Meerovitch KS, Svitkin YV, Lee HS,
Lejbkowicz F, Kenan DJ, et al. 1993.
J. Virol. 67:3798 – 807
51. Alspaugh MA, Talal N, Tan EM. 1976.
Arthritis Rheum. 19:216 –22
52. Akizuki M, Powers R, Holman HR.
1977. J. Clin. Invest. 59:264 –72
53. Habets WJ, den Brok JH, Boerbooms
AM, van de Putte LB, van Venrooij WJ.
1983. EMBO J. 2:1625–31
54. Smith PR, Williams DG, Venables PJ,
Maini RN. 1985. J. Immunol. Methods
77:63–76
55. Rosenblum JS, Pemberton LF, Blobel G.
1997. J. Cell Biol. 139:1655– 61
56. Sobel SG, Wolin SL. 1999. Mol. Biol.
Cell 10:3849 – 62
57. Deng JS, Takasaki Y, Tan EM. 1981.
J. Cell. Biol. 91:654 – 60
58. Graus F, Cordon-Cardo C, Bonfa E,
Elkon KB. 1985. J. Neuroimmunol.
9:307–19
59. Long KS, Cedervall T, Walch-Solimena
C, Noe DA, Huddleston MJ, et al. 2001.
RNA 7:1589 – 602
60. O’Brien CA, Margelot K, Wolin SL.
1993. Proc. Natl. Acad. Sci. USA
90:7250 –54
61. Peek R, Pruijn GJM, van der Kemp A,
van Venrooij WJ. 1993. J. Cell Sci. 106:
929 –35
62. Simons FH, Broers FJ, van Venrooij WJ,
Pruijn GJ. 1996. Exp. Cell Res. 224:
224 –36
63. Shiroki K, Isoyama T, Kuge S, Ishii T,

Ohmi S, et al. 1999. J. Virol. 73:2193–
200
64. Casciola-Rosen LA, Anhalt G, Rosen A.
1994. J. Exp. Med. 179:1317–30
65. Rutjes SA, Utz PJ, van der Heijden A,
Broekhuis C, van Venrooij WJ, Pruijn
GJ. 1999. Cell Death Differ. 6:976 – 86
66. Ayukawa K, Taniguchi S, Masumoto J,
Hashimoto S, Sarvotham H, et al. 2000.
J. Biol. Chem. 275:34465–70
67. Bachmann M, Pfeifer K, Schroder HC,
Muller WE. 1989. Mol. Cell. Biochem.
85:103–14
68. Borer RA, Lehner CF, Eppenberger HM,
Nigg EA. 1989. Cell 56:379 –90
69. Chauvet S, Maurel-Zaffran C, Miassod
R, Jullien N, Pradel J, Aragnol D. 2000.
Dev. Dynam. 218:401–13
70. Tan Q, Li X, Sadhale PP, Miyao T,
Woychik NA. 2000. Mol. Cell. Biol.
20:8124 –33
71. Bandziulis RJ, Swanson MS, Dreyfuss
G. 1989. Genes Dev. 3:431–37
72. Query CC, Bentley RC, Keene JD. 1989.
Cell 57:89 –101
73. Chan EKL, Francoeur AM, Tan EM.
1986. J. Immunol. 136:3744 – 49
73a. Rosenblum JS, Pemberton LF, Bonifaci
N, Blobel G. 1998. J. Cell Biol. 143:
887–99
74. Kenan DJ. 1995. RNA recognition by the
human La protein and its relevance to
transcription, translation, and viral
infectivity. PhD thesis. Duke Univ.
75. Maraia RJ, Intine RV. 2001. Mol. Cell.
Biol. 21:367–79
76. Ohndorf UM, Steegborn C, Knijff R,
Sondermann P. 2001. J. Biol. Chem. 276:
27188 –96
77. Varani G, Nagai K. 1998. Annu. Rev.
Biophys. Biomol. Struct. 27:407– 45
78. Aigner S, Lingner J, Goodrich KJ,
Grosshans CA, Shevchenko A, et al.
2000. EMBO J. 19:6230 –39
78a. Nagase T, Ishikawa K, Suyama M,
Kikuno R, Miyajima N, et al. 1998. DNA
Res. 5:277– 86
78b. Adams MD, Celniker SE, Holt RA,
THE LA PROTEIN 401
Evans CA, Gocayne JD, et al. 2000.
Science 287:2185–95
78c. Matsui Y, Toh EA. 1992. Mol. Cell.
Biol. 12:5690 –99
79. Birney E, Kumar S, Krainer AR. 1993.
Nucleic Acids Res. 21:5803–16
80. Walker JE, Saraste M, Runswick MJ,
Gay NJ. 1982. EMBO J. 1:945–51
81. Craig AW, Svitkin YV, Lee HS,
Belsham GJ, Sonenberg N. 1997. Mol.
Cell. Biol. 17:163– 69
82. Pizer LI, Deng J-S, Stenberg RM, Tan
EM. 1983. Mol. Cell. Biol. 3:1235– 45
83. Francoeur AM, Chan EKL, Garrels JI,
Mathews MB. 1985. Mol. Cell. Biol.
5:586 –90
84. Broekhuis CH, Neubauer G, van der
Heijden A, Mann M, Proud CG, et al.
2000. Biochemistry 39:3023–33
85. Kagami M, Toh-e A, Matsui Y. 1997.
Genetics 147:1003–16
86. Tsukada M, Gallwitz D. 1996. J. Cell
Sci. 109:2471– 81
87. Yu W, Farrell RA, Stillman DJ, Winge
DR. 1996. Mol. Cell. Biol. 16:2464 –72
88. Chang YN, Kenan DJ, Keene JD, Gatig-
nol A, Jeang KT. 1994. J. Virol.
68:7008 –20
89. Svitkin YV, Pause A, Sonenberg N.
1994. J. Virol. 68:7001–7
90. Svitkin YV, Meerovitch K, Lee HS,
Dholakia JN, Kenan DJ. 1994. J. Virol.
68:1544 –50
91. Ali N, Siddiqui A. 1997. Proc. Natl.
Acad. Sci. USA 94:2249 –54
92. Pellizzoni L, Cardinali B, Lin-Marq N,
Mercanti D, Pierandrei-Amaldi P. 1996.
J. Mol. Biol. 259:904 –15
93. Ali N, Pruijn GJ, Kenan DJ, Keene JD,
Siddiqui A. 2000. J. Biol. Chem. 275:
27531– 40
94. Crosio C, Boyl PP, Loreni F, Pierandrei-
Amaldi P, Amaldi F. 2000. Nucleic
Acids Res. 28:2927–34
95. Holcik M, Korneluk RG. 2000. Mol.
Cell. Biol. 20:4648 –57
96. Evans CF, Engelke DR. 1990. Methods
Enzymol. 181:439 –50
402 WOLIN y CEDERVALL
97. Lee Y, Kindelberger DW, Lee JY,
McClennen S, Chamberlain J, Engelke
DR. 1997. RNA 3:175– 85
98. Anderson J, Phan L, Cuesta R, Carlson
BA, Pak M, et al. 1998. Genes Dev.
12:3650 – 62
99. Anderson J, Phan L, Hinnebusch AG.
2000. Proc. Natl. Acad. Sci. USA
97:5173–78
100. Chanfreau G, Elela SA, Ares M Jr, Guth-
rie C. 1997. Genes Dev. 11:2741–51
101. Elela SA, Ares M Jr. 1998. EMBO J.
17:3738 – 46
102. Allmang C, Kufel J, Chanfreau G,
Mitchell P, Petfalski E, Tollervey D.
1999. EMBO J. 18:5399 – 410
103. Seipelt RL, Zheng B, Asuru A, Rymond
BC. 1999. Nucleic Acids Res. 27:587–95
104. Pannone BK, Kim SD, Noe DA, Wolin
SL. 2001. Genetics 158:187–96
105. Will CL, Luhrmann R. 2001. Curr. Opin.
Cell Biol. 13:290 –301
106. Rymond BC. 1993. Proc. Natl. Acad.
Sci. USA 90:848 –52
107. Roy J, Zheng B, Rymond BC, Woolford
JL. 1995. Mol. Cell. Biol. 15:445–55
108. Bordonne R, Tarassov I. 1996. Gene
176:111–17
109. Achsel T, Brahms H, Kastner B, Bachi
A, Wilm M, Luhrmann R. 1999. EMBO
J. 18:5789 – 802
110. Mayes AE, Verdone L, Legrain P, Beggs
JD. 1999. EMBO J. 18:4321–31
111. Salgado-Garrido J, Bragado-Nilsson E,
Kandels-Lewis SB Sr. 1999. EMBO J.
18:3451– 62
112. Vidal VP, Verdone L, Mayes AE, Beggs
JD. 1999. RNA 5:1470 – 81
113. Terns MP, Lund E, Dahlberg JE. 1992.
Mol. Cell. Biol. 12:3032– 40
114. Lund E, Dahlberg JE. 1992. Science 255:
327–30
115. Madore SJ, Wieben ED, Pederson T.
1984. J. Biol. Chem. 259:1929 –33
116. Wolin SL, Steitz JA. 1983. Cell
32:735– 44
117. Simons FHM, Pruijn GJM, van Venrooij
WJ. 1994. J. Cell Biol. 125:981– 88
118. Howe JG, Shu MD. 1988. J. Virol.
62:2790 –98
119. Bertrand E, Houser-Scott F, Kendall A,
Singer RH, Engelke DR. 1998. Genes
Dev. 12:2463– 68
120. Sarkar S, Hopper AK. 1998. Mol. Biol.
Cell 9:3041–55
121. Grosshans H, Hurt E, Simos G. 2000.
Genes Dev. 14:830 – 40
122. Maraia RJ, Sasaki-Tozawa N, Driscoll
CT, Green ED, Darlington GJ. 1994.
Nucleic Acids Res. 22:3045–52
123. Goodier JL, Fan H, Maraia RJ. 1997.
Mol. Cell. Biol. 17:5823–32
124. Cozzarelli NR, Gerrard SP, Schlissel M,
Brown DD, Bogenhagen DF. 1983. Cell
34:829 –35
125. Watson JB, Changler DW, Gralla JD.
1984. Nucleic Acids Res. 12:5369 – 84
126. Campbell FE, Setzer DR. 1992. Mol.
Cell. Biol. 12:2260 –72
127. Belsham GJ, Sonenberg N. 2000. Trends
Microbiol. 8:330 –35
128. Troster H, Metzger TE, Semsei I,
Schwemmle M, Winterpacht A, et al.
1994. J. Exp. Med. 180:2059 – 67
129. Carter MS, Sarnow P. 2000. J. Biol.
Chem. 275:28301–7
130. Parkin NT, Cohen EA, Darveau A,
Rosen C, Haseltine W, Sonenberg N.
1988. EMBO J. 7:2831–37
131. Jackson RJ, Kaminski A. 1995. RNA
1:985–1000
132. Hunt SL, Jackson RJ. 1999. RNA
5:344 –59
133. Svitkin YV, Ovchinnikov LP, Dreyfuss
G, Sonenberg N. 1996. EMBO J.
15:7147–55
134. Belsham GJ, Sonenberg N, Svitkin YV.
1995. In Cap-Independent Translation,
ed. P Sarnow, pp. 85–98. Berlin: Spring-
er-Verlag
135. Meyuhas O. 2000. Eur. J. Biochem. 267:
6321–30
136. Pellizzoni L, Lotti F, Rutjes SA, Pieran-
drei-Amaldi P. 1998. J. Mol. Biol. 281:
593– 608

137. Peek R, Pruijn GJM, van Venrooij WJ.
1996. Eur. J. Biochem. 236:649 –55
138. McBratney S, Sarnow P. 1996. Mol.
Cell. Biol. 16:3523–34
139. McLaren RS, Caruccio N, Ross J. 1997.
Mol. Cell. Biol. 17:3028 –36
140. Pardigon N, Strauss JH. 1996. J. Virol.
70:1173– 81
141. Duncan RC, Nakhasi HL. 1997. Gene
201:137– 49
142. Heise T, Guidotti LG, Chisari FV. 1999.
J. Virol. 73:5767–76
143. Spangberg K, Wiklund L, Schwartz S.
2001. J. Gen. Virol. 82:113–20
144. Matthews MB, Francoeur AM. 1984.
Mol. Cell. Biol. 4:1134 – 40
145. Shi H, O’Brien CA, Van Horn DJ, Wolin
SL. 1996. RNA 2:769 – 84
146. Pruijn GJM, Slobbe RL, van Venrooij
WJ. 1991. Nucleic Acids Res.
19:5173– 80
View publication stats
THE LA PROTEIN 403
147. Samuels ME, Bopp D, Colvin RA,
Roscigno RF, Garcia-Blanco MA,
Schedl P. 1994. Mol. Cell Biol.
14:4975–90
148. Kuhn U, Pieler T. 1996. J. Mol. Biol.
256:20 –30
149. Samuels M, Deshpande G, Schedl P.
1998. Nucleic Acids Res. 26:2625–37
150. Shahied L, Braswell EH, LeStourgeon
WM, Krezel AM. 2001. J. Mol. Biol.
305:817–28
151. Pederson T. 1998. Nucleic Acids Res.
26:3871–76
152. Maraia RJ. 2001. J. Cell Biol.
153:F13–18
153. Tenenbaum SA, Carson CC, Lager PJ,
Keene JD. 2000. Proc. Natl. Acad. Sci.
USA 97:14085–90
154. Wassarman KM, Repoila F, Rosenow C,
Storz G, Gottesman S. 2001. Genes Dev.
15:1637–51