Welcome to Scribd, the world's digital library. Read, publish, and share books and documents. See more ➡
Download
Standard view
Full view
of .
Add note
Save to My Library
Sync to mobile
Look up keyword
Like this
26Activity
×
0 of .
Results for:
No results containing your search query
P. 1
On FlashPaper

On FlashPaper

Ratings:

4.94

(124)
|Views: 157,625|Likes:
Published by Doc
Scribd blog entry #2!
Scribd blog entry #2!

More info:

Published by: Doc on Jan 05, 2007
Copyright:Attribution Non-commercial

Availability:

Read on Scribd mobile: iPhone, iPad and Android.
download as PDF, DOC, TXT or read online from Scribd
See More
See less

12/21/2012

pdf

text

original

 
On FlashPaper
A Format for Sharing Documents on the Web
An image embedded in FlashPaper. (Coffee with a drop of milk)
One of our goals at Scribd is to create "the best way to publish a documentonline." For short and simple documents, it is generally best to display the text inHTML format. However, for any document that is more than a few pages long or has complex formatting, HTML doesn't work very well. We could allow people to just attach their document as a PDF or Microsoft Word file, but then the sitewouldn't be any fun to browse, because users would have to download eachdocument they want to view. We saw the difficulty with sharing formatteddocuments as a real problem, and we looked for a way to solve it.What we found was a product called Macromedia FlashPaper, which we hadactually never heard of. The document you’re reading now, by the way, is inFlashPaper format. FlashPaper documents are Flash SWF files that look andfunction a lot like PDF files.
 
T
FlashPaper works great for technical articles, faithfully preserving formatting.
A
 
role
 
for 
 
the
 
anaphase-promoting
 
complex
 
inhibitor 
Emi2 XErp1,
 
a
 
homolog
 
of 
 
early
 
mitotic
 
inhibitor 
 
1,
in
 
cytostatic
 
factor 
 
arrest
 
of 
 
Xenopus
 
eggs
Jeffrey
 
J.
 
Tung*
, David V.
Hansen*
, Kenneth H.
Ban*
, Alexander V.
Loktev*,
 
Matthew
 
K.
 
Summers*,
John
 
R.
Adler 
III*,
and
Peter 
 
K.
 
Jackson*
†‡
*Department
 
of 
 
Pathology
 
and
 
Program
 
in
 
Cancer 
 
Biology,
 
Stanford
 
University
 
School
 
of 
 
Medicine,
300 Pasteur 
Drive,
 
Stanford,
 
CA
94305
Communicated
 
by
 
Marc
 
W.
 
Kirschner,
 
Harvard
 
Medical
School,
Boston,
 
MA,
 
February
9, 2005
(received
 
for 
 
review
 
November 
30, 2004)
Unfertilized
 
vertebrate
 
eggs
are
arrested
in
metaphase
 
of 
 
meiosis
II
with
high
cyclin
 
B
 
Cdc2
activity
to
 
prevent
 
parthenogenesis.
Until fertilization,
 
exit
 
from
 
metaphase
 
is
 
blocked
by an activity
called cytostatic
factor 
(CSF),
which
stabilizes
 
cyclin
 
B
by
inhibiting
 
the
anaphase-promoting
 
complex
 
(APC)
 
ubiquitin
ligase.
 
The
 
APC
in-
hibitor 
early mitotic
inhibitor 
1
(Emi1)
 
wasrecently
 
found
 
to
be required
for 
 
maintenance
 
of 
 
CSF
 
arrest.
 
We
show here
that
exog-
enous
 
Emi1
 
is
 
unstable
in
CSF-arrested
 Xenopus
 
eggs
and
is
destroyed
by
the
 
SCF
 
TrCP
 
ubiquitin
 
ligase,suggesting
 
that
endog-
enous
 
Emi1,
an
apparent
44-kDa
protein,
 
requires
a
stabilizing
factor.
However,
 
anti-Emi1antibodies
 
crossreact
 
with
 
native
 
Emi2Erp1
 
FBXO43,
a homolog
of 
 
Emi1
and
conserved
 
APC
inhibitor.
Emi2
 
is
 
stable
in
CSF-arrested
 
eggs,
 
is
 
sufficient
 
to
prevent
 
CSF
release,
and
is
rapidly degraded in a
Polo-like
 
kinase
1-dependent
manner in
response
 
to
 
calcium-mediated
eggactivation.
These results
 
identify
 
Emi2
 
as
a
candidate
 
CSF
maintenance
 
protein.
cyclin
 
B
meiosis
maturation-promoting
 
factor 
 
oocyte
 
maturation
o
 
 prevent
 
 parthenogene
sis,
 
unfertilized
 
eggs
 
rom
 
many
an
imals
 
arre
st
 
in
 
met
aphase
 
of 
 
meiosis
 
II
 
(MII).
 
Sperm
 penetration
 
triggers
 
the
 
release
 
rom
 
met
aphase
 
arre
st
 
and
 
the
c
ommencement
 
of 
 
alternating
 
cycles
 
o
 
DNA
 
replication
 
and
 
cell
div
ision
 
in
 
the
 
embr 
 
yo.
 
The
 
regulator 
 
y
 
 basis
 
for 
 
met
aphase
II
arre
st
 
was
 
first
 
characterized
 
in
 
rog
 
eggs
30
years
ago
 
and
ter 
med
 
c
y
tost
atic
 
factor 
 
(CSF)
 
(1).
 
CSF
 
is
 
operationallydefined as
 
an
 
activ
it
y,
 
rather 
 
than
 
a
 
single
 
molecule,
 
 pre
sentin
 
unfer- tilized
 
eggs
 
that
 
 blocks
 
cleavage
 
of 
 
div
iding
 blastomere
s
 
upon
injection
 
(rev
iewed
 
in
 
ref.
 
2).
 
Mos,
 
an
activator 
 
of 
 
the
 
mitogen- activated
 
 protein
 
inase
 
Rsk 
 
 pathway,
is
 
a
 
key
 
c
omponent
 
of 
CSF
 
that
 
appears
 
at
 
the
 
onset
 
of 
 
meiosis
I
 
(MI)
 
and
 
activate
s
 
CSF
to
 
 block 
 
cleavage
 
of 
 
 blastomere
s
 
(3).
The
 
anaphase-promoting
 
c
omplex
 
(A
 
PC)
 
is
 
an
 
E3
 
ubiquitin
ligase
 
that
 
triggers
 
M-phase
 
ex
it
 
 by
 
directing proteasome-
dependent
 
cyclin
 
B
 
destr 
uction
 
(4),
 
re
sulting
 
in
the
 
sw
if 
t
 
inac-
tivation
 
of 
 
the
 
cyclin
 
B
 
Cdc2
 
inase,
 
or maturation-
 
 promoting factor 
 
(MPF)
 
(5,
 
6).
 
A
 
rise
 
inintracellular 
 
calcium
 
af 
ter 
 
fertil- ization
 
induce
s
 
met
aphase
 
II
release
 
 by
 
relieving
 
the
 
A
 
PC
 
rom
repre
ssion.
 
Early
 
mitoticinhibitor 
1
(Emi1),
 
originally
 
cloned
rom
 
a
 
 Xenopus
 
ooc
y
t
e
cDNA
 
librar 
 
y,
 
 blocks
 
the
 
cleavage
 
of 
injected
 
 blastomere
s
similar 
 
to
 
CSF
 
(7)
 
and
 
ef 
ficiently
 
inhibits
the
 
A
 
PC
 
in
 
vitro
 
(8).Recently,
 
Emi1
 
was
 
shown
 
to
 
 be
 
requiredUpon
 
fertilization
 
of 
 
 Xenopus
 
eggs,
calcium
 
signaling
 
i
nactivate
s
CSF
 
arre
st,
 
which
 
require
s
 
the
 
 Xenopus
 
Polo-like
 
inase
1
(Plx1).
The
 
t
a
rget
 
of 
 
Plx1
 
in
 
this
 
 pathway
 
remains
 
unk 
nown
 
(13).
 
I
n
humansomatic
 
cells,
 
MPF
 
and
 
human
 
Polo-like
 
inase
1
(
Plk1)
 
t
arget
Emi1
 
or 
 
deg
radation
 
 by
 
the
 
Skpl
 
Cullin
 
F-box
 protein
 
(SCF)
 
TrCP
ubiquitin
 
ligase
 
(14
 
 –17).
 
Specifically,
 
Plk1
 phosphor 
ylate
s
 
Emi1
 
on
its
 
DSGxxS
 
sequence,
 
creating
 
a
c
onsensus
 
deg
ron
 
recognized
 
 by
TrCP
 
(17).
 
Thus,
 
 Xenopus
 
Emi1(xEmi1)
 
c
ould
 
 b
e
 
a
 
Plx1
 
t
arget
downstream
 
of 
 
calcium
 
signaling.
A
n
 
apparent
 
 paradox
 
is
 
how
 
Emi1
levels
 
are
 
sustained
 
in
 
the
CSF-arrested
 
egg
 
amid
 
h
igh
 
MPF
 
and
Plx1
 
activ
i
tie
s
.
I
n
 
line
 
w
iththis
 
 paradox,
 
a
 
recent
 
report
 
suggests
 
that
Emi1
 
is
 
unstable
 
andundetect
able
 
i
n
 
 Xenopus
 
eggs
 
(18).
 
On
 
the
other 
 
hand,
 
Emi1
appears
 
to
 
 be
 
 pre
sent
 
in
 
mouse
 
eggs
 
(10).
 
In
 
this
study,
we
want
to
 
clarif 
y
 
our 
 
underst
anding
 
of 
 
Emi1
 
regulation
 
in
 Xenopus
 
eggs
and
 
find
 
t
hat
 
Emi2,
 
an
 
Emi1
 
homolog,
 
may
 
c
ontribute
to
 
CSF
arre
st.
Methods
Reagents.
 
Sera
 
rom
 
four 
 
rabbits
 
immunized
 
w
i
th
 
maltose
 
 binding
 protein
 
(MBP)-Emi1
 
fusion
 
 protein
 
were
 
af 
fin
it
y-purified
 
 b
y
 
low-
ing
 
over 
 
a
 
c
olumn
 
of 
 
GST-Emi1
 
i
mmobilized
 
on
 
CNBr-Sepharose
re
sin
 
w
ith
 
acid
 
e
lution.
 
Other 
 
antibodie
s
 
used
 
wereagainst
 
-cate-
n
i
n,
 
cyclin
 
B2,
 
Plx1,
 
Plk1
 
(Zymed),
 
myc
 
epitope,
and
 
actin
 
(Sant
a Cr 
uz
 
Biotechnolog
y).
 
xEmi2
 
was
 
PCR-cloned
om
 
an
 
ooc
y
t
e
cDNA
 
librar 
y,
 
and
 
a
 
human
 
Emi2
 
(
hEmi2)clone
 
was
 
 purchased
rom
 
Inv
itrogen.
 
 pCS2-cDNA
 
c
onstr 
uctswere
 
linearized
 
and
 
in
 
vitro
-transcribed
 
to
 
generate
 
mRNA
 
 by
using
 
a
 
mMe
ssage
 
Machine
i
t
 
(Ambion,
 
Austin,
 
TX).
 
 pCS2-cDNA
 
c
onstr 
ucts
 
were
 
in
 
vitro
-
translated
 
(I
VT)
 
in
 
rabbitreticuloc
y
t
e
 
l
ysate
 
(TNT,
 
Promega)
 
and
labeled
 
w
i
th
 
35
S-methionine.
 
A
ll
 
Emi1
 
and
 
Emi2
 
e
x
 periments
 
used
 Xenopus
sequence
s
 
unle
ss
 
other 
w
i
se
 
noted
 
as
 
hEmi1
 
and
 
hEmi2
 
or 
humansequence
s.
 
MBP-fusion
 
 proteins
 
and
 
GST-Plk1
 
were
 
ex-
 pre
ssed
 
in
 
Escher 
ichia
 
coli 
 
and
 
 purified
 
 b
y
 
 batch
 
 binding
 
 bacterial
 protein
 
l
ysate
 
to
 
af 
fin
it
y
 
e
s
in
 
and
 
elution
 
w
ith
 
maltose
 
or 
gluta-thione,
 
then
 
dialyzed
 
into
 
XB
 
 buf fer 
 
(
20
mM
 
Hepe
s,
 
 pH
7.7 100
mM
 
KCl).
 
Point
 
mut
ations
 
were
 
engineered
 
w
i
th
 
a
QuikChange
 
i
t
(Strat
agene).
Handling
 
of 
 
Xenopus
 
Oocytes.
 
Ooc
y
t
e
s
 
were
 
obt
ained
 
and
 proce
ssed
for 
 
H1
 
i
nase
 
activ
it
y
 
and
 
immunoblot
 
as
 
de
scribed(19).
 
Ooc
y
t
e
s
were
 
injected
 
w
i
th
30
ng
 
of 
 
MBP-Emi1
 
usion
 protein
 
o
10
n
g
 
o
various
 
mRNA
 
in
 
tot
al
 
volume
s
 
not
exceeding
50
nl.
 
Maturation
for 
 
maintenance
 
of 
 
CSF
 
arre
st
 
in
 
rog
 
and
 
mouse
eggs.
Immu-
 
nodepletion
 
of 
 
Emi1
 
rom
 
 Xenopus
 
CSF
 
egg
 
extract
 
cause
s
 
rapid
cyclin
 
B
 
 proteolysis
 
and
 
ex
it
 
rom
 
met
aphase
 
arre
st
 
independent
of 
 
calcium
 
mobilization,
 
and
 
ablation
 
of 
 
Emi1
 
 by
 
small
 
inter 
fer-
ing
 
RNA
 
in
 
mouse
 
ooc
y
t
e
s
 
induce
s
 
 parthenogene
sis
 
(9,
 
10).
Recent
 
work 
 
has
 
shown
 
that
 
the
 
Mos
 
mitogen-activated
 
 protein
inase
 
Rsk 
 
 pathway
 
e
s
t
ablishe
s,
 
 but
 
is
 
not
 
required
 
to
 
maintain,CSF
 
arre
st
 
(11,
 
12).
 
Therefore,
 
CSF
 
arre
st
 
is
 
a
 
c
omplex
 
 proce
ss
e
s
t
ablished
 
 by
 
the
 
mitogen-activated
 
 protein
 
inase
 
 pathway
and
maintained
 
through
 
inhibition
 
of 
 
the
 
A
 
PC.
Freely
available
 
online
 
through
 
the
 
PNAS
 
open
 
access
 
option.
Abbreviations:
 
APC,
 
anaphase-promoting
 
complex;
 
CHX,
 
cycloheximide;
 
CSF,
cytostatic
factor;
Emi,
early
 
mitotic
 
inhibitor;
 
hEmi,
 
human
Emi; xEmi,
 Xenopus
 
Emi;
GVBD,
 
germinal
vesicle
 
breakdown;
 
IVT,
 
in
 
vitro-
translated;
 
MI,
 
m
eiosis
I;
MII,
meiosisII;
MBP,
 
Maltose
binding
 
protein;
 
MPF,
 
mitosis-promoting
 
factor;
Plk1,
human
 
Polo-like
 
kinase
1;
Plx1,
 Xenopus
 
Polo-like
 
kinase
1;
SCF,
Skpl
Cullin
 
F-box
 
protein.
Data
 
deposition:
The
sequence
 
reported
 
in
 
this
 
paper 
 
has
 
been
 
deposited
 
in
 
the
 
GenBank
database
 
(accession
 
no.
 
AY928267).
To
whom
 
correspondence
 
should
 
be
 
addressed.
 
E-mail:
 
pjackson@stanford.edu.
©
2005
by
The
National
 
Academy
 
o
 
Sciences
 
of 
 
the
 
USA
 
4318
 –
4323
PNAS
March 22, 2005
vol.
102
no.
12
www.pnas.org
 
cgi
 
doi
10.1073
pnas.0501108102

Activity (26)

You've already reviewed this. Edit your review.
1 hundred reads
1 thousand reads
1 hundred thousand reads
Dyna P Hd liked this
Sara Keats liked this
dzull06 liked this

You're Reading a Free Preview

Download
/*********** DO NOT ALTER ANYTHING BELOW THIS LINE ! ************/ var s_code=s.t();if(s_code)document.write(s_code)//-->