Plant Cell Rep (2010) 29:419–436 DOI 10.

1007/s00299-010-0826-8

REVIEW

The superfamily of thaumatin-like proteins: its origin, evolution, and expression towards biological function
Jun-Jun Liu • Rona Sturrock Abul K. M. Ekramoddoullah

Received: 7 January 2010 / Revised: 26 January 2010 / Accepted: 28 January 2010 / Published online: 5 March 2010 Ó Her Majesty the Queen in Rights of Canada 2010

Abstract Thaumatin-like proteins (TLPs) are the products of a large, highly complex gene family involved in host defence and a wide range of developmental processes in fungi, plants, and animals. Despite their dramatic diversification in organisms, TLPs appear to have originated in early eukaryotes and share a well-defined TLP domain. Nonetheless, determination of the roles of individual members of the TLP superfamily remains largely undone. This review summarizes recent advances made in elucidating the varied TLP activities related to host resistance to pathogens and other physiological processes. Also discussed is the current state of knowledge on the origins and types of TLPs, regulation of gene expression, and potential biotechnological applications for TLPs. Keywords Antifungal activity Á Defence response Á Environmental stress Á Phylogenetic analysis Á Thaumatin-like protein Abbreviations AFP Anti-freeze protein GPCRs G protein-coupled receptors MYA Million years ago OLP Osmotin-like protein

PR PR5 PR5K TLP

Pathogenesis-related Family 5 of pathogenesis-related proteins PR5-like kinase Thaumatin-like protein

Introduction Thaumatin-like proteins (TLPs) are polypeptides of about 200 amino acid residues that share sequence similarity with thaumatin (Velazhahan et al. 1999), a sweet-tasting protein originally found in the fruit of the West African rain forest shrub Thaumatococcus daniellii (Benth.) (Van der Wel and Loewe 1972). Due to their inducible expression by stresses like pathogen/pest attack, plant TLPs are classified as the pathogenesis-related (PR) protein family 5 (PR5), 1 of 17 families of defence-related PR proteins (Christensen et al. 2002; van Loon et al. 2006). The most recent review of plant TLPs by Velazhahan et al. (1999) describes occurrence, properties, and regulation of plant PR5 genes. Over the past decade, TLPs have been discovered in a wide range of organisms (Shatters et al. 2006), including nematodes (Kitajima and Sato 1999), insects (Brandazza et al. 2004), fungi (Grenier et al. 2000; Sakamoto et al. 2006), and both gymnosperms (Midoro-Horiuti et al. 2000; Piggott et al. 2004; Zamani et al. 2004; Futamura et al. 2006; O’leary et al. 2007; Liu et al. 2010) and angiosperms (Velazhahan et al. 1999; van Loon et al. 2006). This paper provides an overview of the knowledge gained on the TLP superfamily over the last decade. Included are results about the origin, structure, evolution, expression, and function of TLPs as well as new information on their potential biotechnological applications.

Communicated by R. Reski. J.-J. Liu (&) Á R. Sturrock Á A. K. M. Ekramoddoullah Natural Resources Canada, Canadian Forest Service, Pacific Forestry Centre, 506 West Burnside Road, Victoria, BC V8Z 1M5, Canada e-mail: Jun-Jun.Liu@NRCan-RNCan.gc.ca

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Greenstein et al. These food TLP allergens include cherry (Prunus avium L. have also been named permatins. 10 in western white pine (Pinus monticola Dougl. Anzlovar and Dermastia 2003). 2002. Osmotin is a TLP that was originally regarded as a saltinduced protein in osmotically stressed tobacco (Nicotiana tabacum L. Based on available plant genomes and EST databases.) cells (Singh et al.). 2006).) (Fujimura et al. (2000) first reported the presence of TLPs in several basidiomycete fungi.G.) (Roberts and Selitrennikoff 1990. Anzlovar and Dermastia 2003). More recently. 28 in Arabidopsis thaliana (L. 1989).) and Rhizoctonia solani (J. and Orthoptera (Schistocerea) (Shatters et al. 2006). ex D. 1999. and PR5-like kinase receptors TLPs are universal in plants.). and found that fungi TLP family is small. 2006). 2004). 2003). hordomatin from barley (Hordeum vulgare L. Several members of the plant TLP superfamily have been reported as food allergens from fruits and pollen allergens from conifers (Hoffmann-Sommergruber 2002. and a total of three in L. 2006). Animal TLPs Animal TLPs were first reported in the nematode Caenorhabditis elegans (Maupas) (Kitajima and Sato 1999). Hemiptera (Toxoptera). f. JGI). Patens (Table 1). Although a few previous reports considered cet A and its ascomycete homologs as TLPs (Sakamoto et al. Cup a3 from Arizona cypress (Cupressus arizonica Greene) (Cortegano et al. but no TLP gene was retrieved from EST data of the single-celled green alga (Chlamydomonas reinhardtii P. ¨ Kuhn). Aspergillus nidulans cet A showed certain homology with S-type TLPs with molecular masses of 16–17 kD from cereal plants (Osherov et al. 2003). van Loon et al.). 2010). Hymenoptera (Lysiphlebus). Arabidopsis and rice both contain PR5-like receptor kinases (PR5K) with an extracellular TLP domain and an intracellular kinase domain (Wang et al. gymnosperms.) (Liu et al. Our BLAST search through the available genome databank revealed the TLP family represented by six members each in C. 123 . which are believed to be capable of creating transmembrane pores (Abad et al. and banana (Musa acuminate Colla) TLP (Leone et al. To date no TLP has been found in Drosophila. and Pru av2 each show a binding ability to Ig-E from allergic persons by allergenic motifs in their protein structures (Ghosh and Chakrabarti 2008). Arabidopsis contains three PR5K genes while rice has at least one (Table 1). Of pollen TLP allergens. wounding and cold hardiness. fungal TLP-encoding genes have also been found in the basidiomycete yeast Cryptococcus neoformans (Sakamoto et al. and bryophytes like the moss Physcomitrella patens subsp. drought. A. Cryptococcus neoformans. plant TLPs were assigned to form family 5 of the PR proteins (Velazhahan et al. and Cry j3 from Japanese cedar (Cryptomeria japonica L. including pathogen/ pest invasion. A recent study found that TLPs are present at least in four insect orders: Coleoptera (Diaprepes and Biphyllus). osmotin and osmotin-like proteins (OLPs) are basic isoforms and they presumably accumulate in the vacuoles of plant cells instead of extraˇ cellularly (Yun et al.) (Brandazza et al. Don). Breiteneder 2004). Because environmental stresses. and avematin from oats (Avena sativa L. grape (Vitis vinifera L. bell pepper (Capsicum annuum L. apple (Malus x domestica) Mal d2 (Krebitz et al. 2000). PR5-like allergies. 2006). osmotin-like proteins. 13 in white spruce (Picea glauca Moench). PR5K genes are present in both monocots and dicots. Dang. Among these allergenic TLPs. Plant TLPs: PR5. including angiosperms. it is worthwhile to point out that so far no protein with typical TLP-domain features has been found in ascomycete fungi. 2007). Jun a3. As the genome sequences of several fungi are available. we performed BLAST search through the available genome databank of the Gene Index Project (at the Computational Biology and Functional Genomics Laboratory. Lentinula edodes (Berk. Buchholz) (Midoro-Horiuti et al. there are Jun a3 from mountain cedar (Juniperus ashei J. only two or three members in basidiomycota Coprinopsis cinerea. the Dana-Farber Cancer Institute) and Fungal Genome Resources (at the Broad Institute and the DOE Joint Genome Institute. 1996). 2000). Mal d2. migratoria (Table 1). 2002). ´ 2002).) Act c2 (Gavrovic´ Jankulovic et al. 6 in moss (Table 1).).) Pru av2 (Inschlag et al. In contrast to most other plant PR5 proteins. and Moniliophthora perniciosa (Table 1). five in the red flour beetle (Tribolium castaneum Herbst). including Irpex lacteus (Fr. As for ascomycete fungi. 2004). 1998.420 Plant Cell Rep (2010) 29:419–436 Origins of the TLP genes Fungal TLPs Grenier et al. and ˚ later in insects Schistocerca gregaria (Forsskal) and Locusta migratoria (L. Skadsen et al. the TLP superfamily contains at least 31 genes in rice (Oryza sativa). kiwi (Actinidia chinensis Planch. 1998).) Cap a1 (Fuchs et al. Examples of permatins occurring in particularly high concentrations in cereal seeds include zeamatin from maize (Zea mays L. elegans and the pea aphid (Acyrthosiphon pisum). 2006). Some PR5 proteins. induce their expression. ˇ 1996.) TLP (Pastorello et al.

FC369913 Animal Acyrthosiphon pisum (6) XP_001942530. 2009). NP_177708. NP_001067337. XP_001769333. NP_507263 Fungi Basidiomycete Moniliophthora perniciosa (3) XP_002389185. XP_748371 These sequences were searched out by BLAST through the available genomes at GenBank. XP_001942788.2 Ascomycete Aspergillus nidulans (2) XP_660683. XP_002395825 Coprinopsis cinerea (3) XP_001837753. NP_001059476. NP_180445. DR580767. NP_001054348. EX409023. DR555245. Patens (6) XP_001765997. These large (L-) type TLPs contain 16 conserved cysteine residues. NP_001067334. Based on PROSITE. TLPs are known to have a thaumatin family signature (PS00316): G-x-[GF]-x-C-x-T-[GA]-D-C-x(1.Plant Cell Rep (2010) 29:419–436 421 Table 1 Thaumatin-like protein sequences (total number in brackets) and their GenBank accession numbers or gene locus in representative species belonging to the plant. NP_001050832. and fungal kingdoms Plant Arabidopsis thaliana (28) NP_173261. NP_198818. NP_177641. NP_001067331. DR566085. DR552739. XP_975156. EX409554 Pinus monticola (10) GQ329659 to GQ329668 Physcomitrella patens subsp. NP_001031064. NP_192902. CNAG_05924. and through the Gene Index Project data sets at DanaFarber Cancer Institute. XP_680888 Aspergillus fumigatus XP_747035. XP_001942572. XP_002395649. NP_177182. NP_180054. grubii H99) (2) Locus: CNAG_02497. XP_001837765 Cryptococcus neoformans (var. XP_969010. animals and basidiomycete fungi share significant homology with thaumatin (Fig. NP_001058421. NP_001067074. XP_975166. NP_001064159. XP_001942718. NP_195834. families and functional sites (Hulo et al. NP_001068535. DR584727. BJ186638. NP_195324. XP_001942779. XP_754650. NP_001064158. XP_001837754. NP_001050833. XP_001784610. NP_001067332. XP_001951906 Tribolium castaneum (5) XP_968724. DR551624. 1). XP_975175 Locusta migratoria (3) CO821337. NP_001050827. DR550004. AJ566726. NP_973870. NP_001063785.3)-C (Jami et al. NP_001064589. animal. DV996551. 2007. NP_177893. NP_001031809. NP_173432. NP_179376. NP_195579. The molecular mass of a small group of small (S-) 123 . NP_194149. DR582629. NP_001049473. NP_198644. NP_173365. NP_001078513 Oryza sativa (31) NP_001041821. NP_001067073. CO849988 Caenorhabditis elegans (6) NP_500747. NP_177642.2.2)[GQ]-x(2. DR573433. NP_197850. NP_001067333. NP_502361. Most TLPs have molecular masses ranging from 21 to 26 kD. NP_502360. NP_001067069. NP_500748. CO821344. NP_001052130. NP_001068554 Picea glauca (13) DR548648. NP_001049533. NP_195325. NP_001062246. NP_177503. NP_193559. NP_001044756. NP_001049531. Tachi et al. NP_001067335. the database of protein domains. NP_568046. NP_001068536. NP_001063786. NP_001067336. NP_177640. and Fungal Genome Resources at the Broad Institute and the DOE Joint Genome Institute (JGI) Protein structure Amino acid sequences for TLPs from plants. NP_502362. 2008).

(XP_001765997). 1 Structural features of the TLP superfamily as shown by amino acid sequence alignment of 15 representative TLPs from a variety of taxa. S-type TLPs have only ten cysteines at conserved positions because of a peptide deletion (Fig. Dashes in the sequences represent single amino acid gaps for best alignment. NP_502360). The TLP family signature (PS00316) in thaumatin.422 Fig. as well as for resistance to protease degradation (Smole et al. G-x-[GF]-x-C-x-T[GA]-D-C-x(1. The disulfide bridges formed by these conserved cysteines help stabilize the molecule and allow for correct folding and high stability under extreme thermal and pH conditions (Fierens et al. basidiomycete fungi Coprinopsis cinerea (XP_001837765) and Moniliophthora perniciosa (XP_002389185). and ascomycete fungi Aspergillus nidulans (XP_660683) and Neosartorya fischeri (XP_001258659).2)-[GQ]-x(2. and other conserved amino acids are indicated by a gray shadow. rice (NP_001049473 and NP_001067331). 2009). 2008). is boxed. conifer Pinus monticola (GQ329659 and GQ329660). These include Thaumatococcus daniellii (thaumatin. AB265690). Arabidopsis (NP_173261 and NP_173365 for TLPs. The conserved cysteine residues are indicated by a black background. 1). and they are responsible for topology and surface electrostatic potential around the cleft proposed to determine TLP specificity to their target receptors or ligands Plant Cell Rep (2010) 29:419–436 type TLPs mainly from conifers and cereals is around 16– 17 kD. Conserved positions of five amino acids are labeled with an asterisk. pea aphid (Acyrthosiphon pisum. moss Physcomitrella patens subsp. nematode (Caenorhabditis elegans. XP_001942779). Both animal and plant TLPs contain an N-terminal signal peptide targeting mature proteins into the secretory pathway. Tobacco osmotin and 123 . and NP_198644 for PR5K).3)C.

2004). These ascomycete sequences shared 32–92% identity within group but only 3–24% between group identities. For example. plant. Thus far. another study indicated that TLPs are grouped into three major clades representing the animal. crystal structures have been determined for seven plant TLPs. Alignment analysis was done using 118 putative TLP sequences selected from ascomycete and basidiomycete fungi. 2005). zeamatin (Batalia et al. 1992). 1999) and osmotin (Min et al. To one side of domain I is domain II made up of several loops.Plant Cell Rep (2010) 29:419–436 423 plant OLPs also have a C-terminal propeptide that is considered to be a determinant for their vacuolar targeting ˇ (Anzlovar and Dermastia 2003). A so-called thaumatin loop that is present within the thaumatin domain II but absent in the antifungal plant PR5 proteins may explain their differences in function (Min et al. 2006) and tomato NP24-I (Ghosh and Chakrabarti 2008). 1). PmTLP-L1 (GQ329659). tobacco PR-5d (Koiwa et al. In addition to a long deletion. or basic nature for binding different ligands/ receptors. modulated and represented by a ribbon diagram. domain I is a lectin-like b-barrel that forms the compact core of a TLP molecule. This acidic cleft is assumed to be relevant to their specific receptor binding for an antifungal activity (Batalia et al. Especially. neutral. The disulfide bridges formed by the conserved cysteines in TLPs are crucial to their characteristic 3D structures. Koiwa et al.org/). and fungal kingdoms. However. In all plant PR5 proteins with known antifungal activity this cleft is acidic because of five highly conserved amino acids (arginine. Group I contains representative sequences from ascomycete fungi exclusively. we conducted a comprehensive search of available genome sequences and EST database (Table 1). and it is far away from all other TLP groups. each containing proteins with relatively high identity. plus animals as represented by nematodes and insects. with the possibility of nine distinct groups (I–IX). 1). In the 3D structures of these plant TLPs. domain III composed of a junction loop and a b-sheet.expasy. ascomycete sequences have other deletions at N-terminal region (Fig. Evolution Evolutionary relationships within the TLP superfamily are not well understood. 1999. respectively (Sakamoto et al. To better decipher the evolution of the TLP superfamily. Because of their low degree of similarity. 2004). plus the cherry allergen Pru Av2 (Dall’Antonia et al. Very similar topological model is predicated by homology modeling and loop insertion for other TLPs from plants (Fig. similar to that in the S-type TLPs from cereals and conifers. Each domain is stabilized by at least one disulfide bridge linked by up to 16 cysteine residues with a conserved spatial distribution throughout the protein (Min et al. 2 Three-dimensional (3D) structure of one Pinus monticola TLP. domain II made up by three a-helical structures plus two b-sheets. and three aspartic acid residues) (Fig. 1996). 1). The topological pattern of the resulting tree indicates that the TLP superfamily is highly divergent. moss. 2004). including thaumatin (Ogata et al. so it is likely that these so-called ascomycete TLPs may have structures that are distinctly different from other TLPs. gymnosperms and angiosperms. The cleft between domains I and II may have an acidic. these ascomycete sequences may represent genes that are nonhomologous with the other eight groups of TLPs and have a Fig. which consists of a small loop. The Deep View Swiss-PdbViewer software package was used to manipulate the PmTLP-L1 3-D model where domain I formed by a b-sandwich with 11 b-sheets. http://swissmodel. 2) and animals (Brandazza et al. a study of phylogenetic and structural relationships by Shatters et al. they share only four of the 16 cysteines typically occurring at conserved positions in most TLPs and they are lack of thaumatin family signature (PS00316) (Fig. The 3D structure of PmTLP-L1 was simulated using the SWISS-MODEL software package (Swiss Institute of Bioinformatics. 1996. 2006). In contrast is the basic cleft region in thaumatin. (2006) suggested that plant PR5 proteins are paraphyletic in angiosperms and related to animal TLPs from insects and nematodes. 2004). 3). glutamic acid. 2004). Min et al. On the other side of domain I is domain III. banana allergen Ba-TLP (Leone et al. and the cleft structure localized between domains I and II 123 . The neighbor joining algorithm was used to construct a phylogenetic tree (Fig. All show similar 3D structures with three domains and a cleft structure between domains I and II (Ghosh and Chakrabarti 2008).

Pph-). suggesting that these ascomycete sequences should not be considered as TLP genes. The scale at the bottom indicates genetic distance proportional to the amino acid substitutions per site Plant Cell Rep (2010) 29:419–436 different evolutionary origin. Acp-). 28 in Arabidopsis (At-). marked as PR5K.424 Fig. 3 Phylogenetic analysis of 118 TLPs by the neighborjoining (NJ) method. 13 in white spruce (Picea glauca. were grouped here based on their TLP domain similarities to other regular TLPs. six in pea aphid (Acyrthosiphon pisum. and Penicillium chrysogenum (Pec). Nematode TLPs constitute a monophyletic clade as group III while all insect TLPs are 123 . TLPs were labeled according to GenBank accession numbers as shown in Table 1 and clustered into nine clusters (I–IX). 10 in western white pine (Pinus monticola. A phylogenetic tree was constructed based on amino acid alignment of TLP sequences by the Clustal W program. Basidiomycete fungi TLPs include three in both Coprinopsis cinerea (Coc-) and Moniliophthora perniciosa (Mop-). and three in locust (Locusta migratoria. Neosartorya fischeri (Nef-).. Aspergillus oryzae (Aso-). Lom-). Trc-). Group II only consists of members exclusively from basidiomycete fungi with 55–80% within group identity and 23–43% identity to other seven TLP groups from plants and animals and all sequences include the 16 conserved Cys residues (Fig. Ascomycete fungi TLPs include two in Aspergillus nidulans (Asn-) and one in Aspergillus fumigatus (Asf-). Pm-). Arabidopsis and rice PR5-like receptor kinases. Plant TLPs include 30 TLPs in rice (Os-). Animal TLPs include six TLPs in nematode (Caenorhabditis elegans. Cae-). and six in moss (Physcomitrella patens subsp. respectively. Pig-). 1). four in red flour beetle (Tribolium castaneum.

The branch topology of the TLP phylogenetic tree indicates that the nine groups may have evolved from multiple rounds of gene duplication. apple (Gao et al. 3). cotton (Gossypium hirsutum cv. but not at all in other floral organs nor in leaves (Sassa et al. Fig. Delta Pine 62) (Wilkinson et al. (2005) found that Arabidopsis AtTLP1 is expressed specifically in the root vascular bundle in response to colonization by non-pathogenic Pseudomonas spp. 2003). In addition to induced expression by various environmental stresses (Velazhahan et al. For examples. there are two subclades (Fig. one of them contains all six moss TLPs that is proposed to have expanded from a single precursor. already possessed at least one TLP gene with typical TLPdomain features (Fig.) TLP family (Futamura et al. these genes were probably already quite diverse in terms of both sequence and function. Kenton et al. 2000). phureja) (Ruiz et al. abscisic acid. VIII. 1996). and rice (Shatters et al. there is additional recent 123 . coordinated with other types of cis-elements. 2010). VII. and methyl jasmonate). Arabidopsis.) TLP gene (PsTL1) was expressed specifically in the pistil and at low level in anthers. In group VII. 2006). and IX). VI. A pepper TLP (PepTLP) was also shown to have a gene expression pattern regulated by fruit-developmental stages (Kim et al. A promoter-gus fusion study found that a Japanese pear (Pyrus serotina Burm. The fact that our phylogeny reconstruction reveals five plant TLP clades containing putative genes from both angiosperm and gymnosperm lineages (Fig. 3). suggesting that group VII may represent an ancestral type of plant TLP superfamily as compared to the other four groups (IV. wounding. wounding and plant hormones (e. VIII. To date. This suggests that tandem duplication be one of the important mechanisms for the asymmetric expansion of the TLP superfamily during species formation. means that a minimum of five TLP genes were probably present in the last common ancestor of extant seed plants about 300 MYA. 1999. which existed about 1 billion years ago (Doolittle et al. Grouping of multiple members of the same species in the same cluster or subcluster suggests that gene duplication events continued to happen throughout the evolution of plant and animal species (Shatters et al. 1997). Jung et al. but has 45–52% identity with angiosperm TLPs in the same cluster (group IV. Characterization of a TLP family in moss strongly suggests the most recent common ancestor of mosses and vascular plants. monticola TLPs (PmTLPs) except for PmTLP-L1 (GQ329659) share an identity threshold of 48% and are grouped in one cluster (group IX. 2005. suggesting that its potential ligand may be different from other PmTLPs and from Plant PR5 protein with known anti-fungal activity (Liu et al. present about 425 million years ago (MYA) (Wellman et al. 2002). are required for the Rtlp1 promoter to respond to fungal elicitors (Hiroyuki and Terauchi 2008). animals and fungi. some TLPs are expressed constitutively in an organ-preferred or development-dependent pattern (Regalado and Ricardo 1996). A recent study of the rice TLP (Rtlp1) found that W-box elements in the promoter region. ´ Leon-Kloosterziel et al. Expression of the TLP superfamily Velazhahan et al. Also. 2000. 2002). For example. all P. 3). but PmTLP-L1. PmTLP-L1 has relatively basic surface in cleft region. GCC boxes. In the tobacco osmotin promoter. Onishi et al. Fig. especially when compared with other PmTLPs with acidic clefts. TLP gene expression switches from the ovary wall to the aleurone (Skadsen et al. These observations suggest that some TLPs may be involved in physiological processes other than host defence to biotic/abiotic stress. 2010). The other subclade of group VII consists of TLPs from both angiosperms and gymnosperms. At that time. 2010). 2002). (1999) reviewed the regulation of plant PR5 gene expression by factors such as microbial infection and pathogenic elicitors. are required for full ethylene responsiveness (Raghothama et al.Plant Cell Rep (2010) 29:419–436 425 localized in another distinct monophyletic clade for group V. (1999). plant TLP genes appear to be confined to multicellular plants (Liu et al. 1). An elderberry (Sambucus nigra) TLP shows a fruit-specific expression and no antifungal and glucanase activities during the final stages of fruit maturation (Van Damme et al. VI. The roles of TLPs and the cascades that regulate their expression differ among members of the Cryptomeria japonica (L. Clustering of multiple TLP genes on the same chromosome and even at the same locus has been observed in a hybrid potato (Solanaceae tuberosum 9 S. f. 3). Most PmTLP members in western white pine are responsive to rust infection. and IX) of plant TLPs. we hypothesize that one TLP gene was present in the last common ancestor of extant plants. 2005). 2006). PmTLP-L1 shows only 30–37% identity to other PmTLPs. 2006). Biological functions Antifungal activity While TLP antifungal activity was well documented by Velazhahan et al. ethylene. 2006).g. the binding site of transcription factor WRKY proteins. salicylate. or exposure to cold temperatures. Plant TLPs are divided into five groups (IV. 2005). in developing barley and oats seed. PmTLP-L5 and PmTLP-S4 show constitutive expression at low levels without significant responsiveness to any tested environmental stresses (Liu et al. f. osmotic stress. Based on this phylogenetic pattern. 2005).

in vitro antifungal activity test.glycines.3-glucans (Trudel et al. members of some fungal TLP families have also been reported to display b-1. For example. Animal TLPs are considered as potential components of the antifungal strategy in animal defence (Brandazza et al. (2007) show that glucanase secreted by some pathogenic fungi and other filamentous fungi can degrade fungal cell walls. parasitica) under in vitro conditions (Campos et al.3-glucanase activity is apparently lacking antifungal activity towards pathogenic fungi (Barre et al. 2007). The recombinant Solanum nigrum TLP (SnOLP) reveals antifungal activity toward several plant pathogenic fungi (Fusarium solani f. 2000. A later study found that only specific barley TLP isoforms interact tightly with b-1. Pr22-3 has glucanase activity. An endo-glucanase from the edible shiitake mushroom Lentinula edodes shares sequence homology with the antifungal TLPs (Grenier et al. Purified zeamatin was 123 . 2000). Martin et al. Grenier et al. Several TLPs exhibit a special binding activity to several water-insoluble b-1. 2003). Mycosphaerella arachidicola. 2007).4-xylanases.3-glucanase activity conferred by plant TLPs is not always related to their anti-fungal activity (van Loon et al. a storage glucan. 2003). Despite the evidence presented above. 2003). it is proposed that xylanase inhibitor activity is one of the mechanisms by which TLPs function in plant defence (Fierens et al. increasing their resistance against pests and pathogens (Selitrennikoff et al. and Physalospora piricola (Chu and Ng 2003). Sakamoto et al. b-1. 2004). acting on vaginal murine candidosis cells. The Kweilin chestnut (Castanopsis chinensis) TLP exerts antifungal activity against Botrytis cinerea. Interestingly. The grapevine TLP (VvTLP-1) significantly inhibits in vitro spore germination and hyphal growth of Elsinoe ampelina (Jayasankar et al. 1999. Franco et al. one of the most abundant polysaccharides (next to cellulose) in the cell wall of higher plants. EC 3.8) depolymerize xylan. Wheat TLXI is among the subgroup of S-type TLPs (15–17 kD). In fact. as indicated by its ability to digest laminarin. Interaction of TLPs with b-1. Fusarium oxysporum. A recent study has discovered that a novel type of xylanase inhibitor (TLX1) from wheat (Triticum aestivum) belongs to the TLP family (Fierens et al.2.) has in vitro antifungal activity against Trichoderma viride and Fusarium oxysporum and shows strong synergistic effects with CsCh1. Some TLPs produced by nematodes and insects are likely to serve to protect them against predation and pathogenic infection by nematophagous and entomopathogenic fungi. Colletotrichum spp.) displays antifungal activity against the snow mold fungus (Microdochium nivale) (Kuwabara et al. TLPs have glucan binding and glucanase activities Mechanisms for the antifungal activity of TLPs are believed to be related to their ability to destroy of the cell walls of pathogenic fungi using a variety of enzymatic activities. an abundant endochitinase from the chestnut cotyledon (Garcia-casado et al. or in transgenic plants. Based on the sequence homology between TLX1 and other TLPs. edodes TLP. 2007).1. and Macrophomina phaseolina) and an oomycete (Phytophthora nicotiana var. 2000). the apple and cherry TLPs possess a moderate endo-b-1. A cold-induced TLP in the apoplast of winter wheat (Triticum aestivum L. identified as TLPs by immunological method and N-terminal sequencing. A banana fruit TLP that exhibits a low but detectable in vitro endo-b1. 2006). Do TLPs function as a-amylase and trypsin inhibitors? Zeamatin is mainly known for its antifungal activity and proposed as a medical agent. 2001).3-glucan may act in concert with PR2 enzyme to disrupt fungal cell wall synthesis and/or prevent proper fungal wall assembly during hyphal extension (Bormann et al. suggesting that the tlg1 endo-glucanase activity helps defend against fungi that are pathogenic on other fungi. providing further evidence to support the suggestion that fungal TLPs are involved in fungal–fungal interactions. 2000). exhibits cell wall lytic activity to degrade lentinan (an antitumor polysaccharide) during postharvest preservation. 1998.3-glucan is a common component of the fungal cell wall. exhibits strong in vitro antifungal activity by blocking the growth of Phomopsis viticola and Botrytis cinerea mycelia (Monteiro et al. 2008). 2002). This enzyme has been shown to be indispensable for the pathogen Botrytis cinerea to infect plants (Brito et al. 2002). The recombinant TLX1 protein displays xylanase-inhibiting activity against Trichoderma longibrachiatum xylanase (XynI) (Fierens et al. 2006).3-D-glucans (Osmond et al. 2002). These findings suggest that glucanase activity is not the sole mechanism for TLP antifungal activity. 2001). b-1. TLPs function as xylanase inhibitor Xylanases (endo-b-1. or related protein activity analyses (Table 2). both plants and microorganisms produce xylanases. sp. Furthermore. tlg1. (2006) revealed that the L. the TLP isolated from chestnut (Castanea sativa Mill. respectively.426 Plant Cell Rep (2010) 29:419–436 evidence supporting the roles of TLPs in host defence and other physiological processes by transgenic overexpression.3-glucanase activity (Grenier et al. 1999. Osmond et al. Also. Two other grape proteins. Pr22-3 was more active in the spore bioassay with pathogens like Rhynchosporium secalis than other barley TLPs (Zareie et al. 2000). 1999.3glucanase activity but are devoid of antifungal activity too (Menu-Bouaouichea et al. Among barley TLPs. 2000).

vulgare Pr22-3 Leaves Mycelium Fruiting bodies Fruiting bodies Fruits Viroid-induced Fruits Fruits. salt or/ and drought Stigmas Callus Endo-b-1. vulgare Leaves Zareie et al. but no endo-b1. (2000) In vitro antifungal activity Vitali et al. Husaini and Abdin (2008). (1999) Kobayashi et al. edodes AP24. tabacum 123 CBP2 N. (1997). basjoo Osmotin N.SE39b N. (2007) Yun et al. vulgare TLP Irpex lacteus TLP Lentinus edodes tlg1 L. 2001) Narasimhan et al.Table 2 A summary of functionally characterized thaumatin-like proteins (TLPs) Regulation/induction (by/in) Constitutively Cultured cells Cotyledon Seed Beetle larvae Chemical in leaves Germinated grain (1. in vitro antifungal activity. MeJa induced in root Fruits Fruits NaCl in cultured cells H.3)-b-glucan-binding activity. (2000) 427 tlp-1 H.3-glucanase activity In vitro antifungal activity (a) Bind to yeast glycoprotein. (2004) Barthakur et al. D’Angeli and Altamura (2007).3-glucanase activity. (2000) Menu-Bouaouichea et al. (2000. (1999) Fagoaga et al. (2006) Transgenic antifungal protection Guo et al.3-glucanase activity Endo-b-1. (2001) Menu-Bouaouichea et al. and antifungal activity Transgenic antifungal protection Endo-b-1.3-glucanase activity. (2003) Barre et al.3-glucanase activity. (d) Transgenic protection to cold. (2005) Salzman et al. and antifungal activity In vitro antifungal activity. (2000) Sakamoto et al.3-glucanase activity.3-glucanase activity. tabacum SE22. (1999) Osmond et al. and antifungal activity Thermal hysteresis Grenier et al. esculentum Mal-TLP Malus domesticus Ban-TLP Musa acuminate Ban-TLP M. but no endo-b1. Parkhi et al.3-glucanase activity Endo-b-1. and in vitro antifungal activity Endo-b-1. acuminata TLP(20kD) M. (2007) Grenier et al. transgenic antifungal protection. but no antifungal activity Endo-b-1. (b) Bind to GPCRs. (2009) Grenier et al. (2000) Grenier et al. (2001). Ibeas et al. (2002) Mackintosh et al. and transgenic antifungal protection Endo-b1. (c) Antifungal activity by glycan interaction.3-glucanase activity. but no in vitro antifungal activity In vitro antifungal activity. (2001) In vitro antifungal activity Chu and Ng (2003) Wang and Duman (2006) In vitro antifungal activity Garcia-Casado et al. (2003) Function/biological role Reference Name Sources PR5 K Arabidopsis thaliana CdTLP Cassia didymobotrya CsTL1 Castanea sativa CaTLP Castanopsis chinensis Dcan1 Dendroides Canadensis Plant Cell Rep (2010) 29:419–436 IFW19 Hordeum vulgare HvPR5c H. tabacum . (2003) Ho et al.3-glucanase activity Bind to cytokinin Endo-b-1.NP24 Lycopersicon esculentum P23 L.3-glucanase activity Endo-b-1. (2006) Grenier et al.

Cold. (2003) In vitro antifungal activity a-amylase/trypsin inhibitor? Monteiro et al. (1999) Menu-Bouaouichea et al. Fu et al.428 Table 2 continued Regulation/induction (by/in) CMV-infected leaves Fungus induced In vivo bind to CMV-1a protein Transgenic antifungal protection Function/biological role Reference Kim et al. (2007) Campos et al.3-glucanase activity. (2003) ´ Richardson et al. (2006). (2002). (1987). (2007) Jayasankar et al. (2002) Fierens et al.3-glucanase activity. tuberosum x demissum Thaumatin II Thaumatococcus daniellii Thaumatin T. tabacum Oryza sativa CHTL Prunus avium Pru-TLP P. avium TLP Rhizoctonia solani TLP(16. dulcamara CBP2 S. salinity and drought protection In vitro antifungal activity Xylanase inhibitor In vitro antifungal activity Name Sources NtTLP1 TLP-D34 N.OLP V. Schestibratov and Dolgov (2005) Li et al. nigrum OLP S.25kD) Secale cereale SniOLP Solanum nigrum SnOLP S. (2003) Grenier et al.3-glucanase activity Antifreeze activity Endo-b-1. Kalpana et al. (2005). daniellii WAS-3a Triticum aestivum TLX1 T. aestivum VVTL-1 Vitis vinifera TLP. (1999). (2007) Grenier et al. (2007) Kuwabara et al. Bind to actin Transgenic antifungal protection Cryoprotection In vitro antifungal activity In vitro antifungal activity.3-glucanase activity. (2005) Datta et al. ABA and elicitors Grains Embryogenic cultures Fungi-infected leaf and berry Grains (b) Transgenic antifungal. vinifera Zeamatin Zea mays Plant Cell Rep (2010) 29:419–436 . (2000) Yu and Griffith (1999) Jami et al. (2002) Rajam et al. Gomez-Leyvaa and Blanco-Labraa (2001) 123 Fruits Fruits Mycelium Cold-induced in leaves Osmotic stress and others – Stems Cultured cells Fruits Fruits (a) Bind to GPCRs. Velazhahan and Muthukrishnan (2003). but no endo-b1. (1997) Maria et al. Maruthasalam et al. (2008) Newton and Duman (2000) Takemoto et al. but no antifungal activity Endo-b-1. and antifungal activity but no antifungal activity Endo-b1.

The binding of osmatin with phosphomannan is required for the maximal TLP toxicity to walled cells but not spheroplasts. alkali-insoluble glucans. human GPCRs are the thaumatin sweet taste receptors (Li et al. and Cassia didymobotrya TLP (CaTLP) showed very different folding characteristics. osmatin induces programmed cell death in yeast (S. (2006) demonstrated that the rough lemon (Citrus jambhiri Lush. These resistance determinants probably form ‘‘barriers’’ that prevent TLP transportation across the cell wall. The microbial cell wall contains resistance determinants to TLPs such as osmotin. but K? blocks this interaction by competing for binding to mannosephosphate groups. genes encoding putative receptor kinases with an exocellular TLP domain (PR5K) have been identified in Arabidopsis (Wang et al. 2005). The yeast mutant (ssd1) is sensitive to osmotin due to its deficiency in cell wall glycoproteins of the PIR (proteins with internal repeats) family. 2003).) miraculin-like protein (RlemMLP2) contains a thaumatin motif and shows both antifungal activity against Alternaria citri and trypsin inhibitor activity. 2000). 2001). Ibeas et al. Narasimhan et al. 2002). TLPs mediate signaling by interaction with their receptors or ligands Most TLPs have specific wide-spectrum antifungal activities. 123 . Phosphomannans are conjugated to several cell wall proteins and several cell wall mannoproteins could bind to immobilized osmotin in vitro (Ibeas et al. Also.) (Richardson et al. Despite having a high structural similarity. suggesting that carbohydrate binding is a common feature of TLPs that determines target specificity. On the other hand. Shiu and Bleecker 2001). the other small band corresponds to a contaminant 14 kD protein that functions as the a-amylase/trypsin inhibitor. 2002). The toxic activity of osmatin on A. cerevisiae) by signaling suppression of stress-responsive genes via the RAS2/cAMP pathway (Narasimhan et al. 1998. 2001). zeamatin. (2000) found that tobacco osmotin binds to phosphomannan. 2005). 2001). 1987. cerevisiae). 2003). thaumatin. respectively (Veronese et al. Salzman et al. Several other TLPs have been shown to bind glucans in vitro (Trudel et al. 2003). (2004) found that Ca2? facilitates binding of osmotin to the fungal cell surface. Tsukuda et al. A 22 kD potato OLP and a 26 kD tobacco OLP were found with binding activities to actin (Takemoto et al. 2000). Selectivity against target microorganisms is probably due to a variety of defense and susceptibility capabilities of the host and target microorganism. 1997. Gomez-Leyvaa and Blanco-Labraa (2001) found that purified zeamatin actually contains five different fractions: four of them correspond to different 22 kD isoforms showing antifungal activity but no inhibitory activity on a-amylase/trypsin. as evidence by a study that found a N. For example. respectively. ´ 2001). 2005). A full understanding of the antifungal mode of action of TLPs is clearly incomplete and still under debate (Franco et al. TLPs do not just interact with fungal cell wall components. This specific in planta interaction with a virus replication-associated protein suggests that NtTLP1 may affect CMV multiplication as well the rate and extent of CMV spreading (Kim et al. 1997) and cytokinin (Kobayashi et al. 1998). cerevisiae involves in a heterotrimeric G protein mediated signal pathway (Yun et al. These results suggest that the GPCR-binding activity may be a conserved characteristic of TLPs (Veronese et al. Coca et al. SchimolerO’Rourke et al. 2000). Overexpression of a heterologous cell wall glycoprotein in Fusarium oxysporum leads to an increased virulence and resistance to osmotin. suggesting that zeamatin’s antifungal activity is not the result of inhibition of these enzymes (Schimoler-O’Rourke et al. nidulans and S. suggesting that phosphomannan promotes osmotin uptake across the cell wall. A receptor-like polypeptide with seven transmembrane domains and structural features of G protein-coupled receptors (GPCRs) is the osmotin binding plasma membrane protein in S. 2003). 1996. Interestingly. Susceptibility of Aspergillus nidulans to osmotin is negatively correlated with cell wall chitin content (Coca et al. 2000). Resistance of the model fungus S. 2003. and other unidentified cell wall components (Yun et al. which indicates that fungal cell wall components can affect resistance to plant defense proteins and pathogenic virulence (Narasimhan et al. zeamatin did not inhibit fungal a-amylase and fungi do not contain trypsin. a cell wall glycoprotein of yeast (S. These authors thus suggest that the protease and a-amylase inhibitory activity ´ should not be assigned to the PR5 proteins (GomezLeyvaa and Blanco-Labraa 2001). Ibeas et al. cerevisiae to osmotin is strongly dependent on the natural polymorphism of its SSD1 gene. tabacum TLP (NtTLP1) interacting with cucumber mosaic virus (CMV) 1a protein in vitro and in planta (Kim et al. cerevisiae that regulates lipid and phosphate metabolism and is required for full sensitivity to osmotin (Narasimhan et al.Plant Cell Rep (2010) 29:419–436 429 able to inhibit porcine pancreatic trypsin and digestive aamylases of insect (Tribolium castaneum) and bacteria (Bacillus sp. The antifungal activity of TLPs also involves in signaling by specific plasma membrane components of the fungal targets. However. suggesting that target recognition may be determined by their interaction with pathogen cell surface components. The fungal cell surface components enhance or suppress TLP antifungal activity (Veronese et al.

It is hypothesized that genes encoding PR proteins arose from gene duplication followed by mutation. Brandazza et al.3-glucanase activities. 2010). Asp97. AFPs (Wang and Duman 2006). 2005). It was found that potato OLP binds to actin directly (Takemoto et al. 2008). and salinity) Freeze-tolerant organisms survive subzero temperatures by the accumulation of anti-freeze proteins (AFPs) that have the ability to adsorb onto the surface of ice crystals and retard ice crystal growth (Griffith and Yaish 2004). fungal cell wall mannoproteins (Ibeas et al. Osmotin overexpression shows multiple effects on programmed cell death.3-glucan binding and b-1. 2004. AFPs accumulate in winter rye leaves in response to cold. drought. located in the central cleft region of the tobacco NP24-I protein are Glu84. resulting in their different forms and functions (Yeh et al. which constitute a catalytic Glu–Asp pair required for the b-1. TLP molecules differ from each other in the distribution of charges on their surface. How have the TLPs adapted diverse biological functions? TLPs appear to have different functions in host–pathogen interaction. and cell signaling. even isoform specific. structures. which contribute to cold protection of the transgenic olive tree (D’Angeli and Altamura 2007). Recently. 2001) do not have antifreeze activity. and/or gene recombination have probably contributed to the origin of novel TLP genes for adaptively important new functions (Wang et al. cytoskeleton organization and Ca2? signaling. 1996. 1997). For example.3-glucan (Trudel et al.430 Plant Cell Rep (2010) 29:419–436 which was considered to affect their roles significantly (Perri et al. Because these interactions are usually family member specific. domain juxtapositions. 2005). they bind to ice crystal directly via hydrophobic interaction and modify the freezing process by inhibiting intercellular ice growth and recrystallization in planta (Griffith et al.3-glucanase activity. drought or ethylene (Yu et al. Role of TLP genes in response to abiotic stresses (cold. stress tolerance. Such findings indicate that the domains of plant PR5 proteins can recognize various self and environmental signal molecules and may be involved in host defence as well as other mechanisms. Natural selection of genes with new functions under environmental pressure has very likely played an important role in the evolution of the genetic diversity of this complex gene family. Asp102. Instead. cytokinin (Kobayashi et al. TLPs from both plants and animals also show a cryoprotection role. including TLPs and other PR proteins (glucanases and chitinases). 2002. indicating that PR proteins with and without antifreeze activity are regulated by different signal transduction pathways. which include actin (Takemoto et al. while crystallographic studies have provided powerful insights into their structural biology. In addition to their roles in pathogen resistance. The PR proteins that accumulate in winter rye leave apoplasts in response to abscisic acid (Yu and Griffith 2001) or salicylic acid (Yu et al. The electronegative character of the interdomain cleft in TLP 3D structure makes an important contribution towards their b-1. Ancestral gene duplication followed by short sequence insertions or deletions.and 25-kD) were revealed to function as AFPs in cold acclimated winter rye (Yu and Griffith 1999).3-glucanase for the hydrolytic cleavage of glycan polymers (Ghosh and Chakrabarti 2008). insertion or deletion of some key amino acid residues during evolution have resulted in the family diversification for potential multiple functions. salt tolerance in transgenic strawberry (Husaini and Abdin 2008) and drought tolerance in transgenic cotton (Parkhi et al. 2002. xylanase (Fierens et al. 2000). A major unanswered question regarding the molecular mechanism underlying TLP biological functions is how the conserved TLPs have adapted functional diversification. 2000). Resolution of this question depends on understanding the interaction of distinct TLPs with a variety of their binding ligands/target proteins. do not act as cryoprotectants. The mechanism for TLPs’ role as cryoprotectant appears complex. 2009). even though TLPDcan1 itself lacks AFP activity (Wang and Duman 2006). In addition to enhancing cold tolerance. 2005). a beetle (Dendroides Canadensis) TLP (TLP-Dcan1) was found to significantly enhance the thermal hysteresis of AFPs by interaction with these proteins. overexpression of the tobacco osmotin gene also improves plant tolerance to salinity and drought stresses in transgenic tobacco (Barthakur et al. viral proteins (Kim et al. Narasimhan et al. while a ´ ˚ longer distance ([5 A) between this catalytic Glu–Asp pair may account for a lower level of its activity as compared to authentic b-1. 1997). Two apoplastic TLPs (16. 2001). suggesting a functional role in freezing tolerance (Newton and Duman 2000). Rep et al. Liu et al. 1998). Winter rye AFPs. Advance of genomics study through the availability of many genome and EST databases (Table 1) has allowed classification of TLPs based on their origins. human or fungal GPCRs (Li et al. Such a plant OLP was purified from Solanum dulcamara stems. 2007). we hypothesize that substitution. and expression profiles. ice crystal (Yu and Griffith 1999). and human Ig-E (Ghosh and Chakrabarti 2008). rather. 3D modeling showed a similar catalytic Glu–Asp pair in the acidic 123 . plant TLPs accumulate in the apoplasts to enable plant adaptation to cold climates. 2000). b1. 2001).

The TLP family thus presents a functional genomics conundrum to answer how it is that a complex protein family can diversify to fulfill multiple functions while conserving their structure of the TLP domain. it has been found that two cold-induced members possess antifreeze activity while other pathogen-induced members have been shown only enzymatic activity (Yeh et al. 2009). Potential application Application of thaumatin as sweetener and flavor enhancer Thaumatin has wide applied usage in food and pharmaceutical industries because it is 1. suggesting that the positive charge of this basic amino acid residue (His22) is crucial for TLX1-xylanase interaction. Application of TLP genes in engineered plant disease resistance In the past decade several transgenic plants have been developed to express a variety of TLP genes for enhanced resistance against pathogens. thaumatin has been approved and commercialized as a safe sweetener and flavor enhancer in food (Zemanek and Wasserman 1995. 1999. why they are so numerous and subject to such complex expression profiles regulated by environmental factors and developmental stages. In the PR3 family of winter rye chitinases. Overexpression of barley TLP-1 in transgenic wheat lines resulted in significant reductions in Fusarium head blight severity in greenhouse evaluations (Mackintosh et al. higher germination percentage.) fruits with thaumatin II accumulation exhibited a sweet phenotype and showed a positive correlation between thaumatin accumulation levels and intensity of sweetness (Maria et al. A similar case has been observed in the PR14 family of plant lipid transfer proteins (LTPs). Faus 2000). Lys106. 2002). 2003). 2003). Since 1983. Thus. specific for thaumatin and not existent in non-sweet TLPs. a site mutagenesis revealed that the mutant TLX1 protein (His22 to Ala22) was unable to form a complex with xylanases. 2007). Lys97. Such findings suggest that amino acid substitutions followed by natural selection may have resulted in adaptive functional alterations for some plant PR proteins. Site-directed mutations in the AFP ice-binding face caused the loss of up to 90% of their antifreeze activity.600 times sweeter than sucrose on a weight to weight basis. Transgenic plants constitutively overexpressing TLPs often show an enhanced fungal resistance (Datta et al. the thaumatin gene was transformed into potato (Witty and Harvey 1990) and tomato (Bartoszewski et al. These five lysine residues. A recent study showed that transgenic expression of the thaumatin II gene resulted not only in a sweeter taste of cucumber fruits in comparison with the control. resulting in a protein lacking xylanase-inhibition capacity (Rombouts et al. and seedling survival under salinity and PEG-mediated drought stress (Rajam et al. Recently. For taste modification of vegetables. 2009). 2001. but also higher aroma acceptability (Zawirska-Wojtasiak et al. and how they interact with their specific ligands? Continued analysis of this protein superfamily will no doubt reveal many other examples of their functional diversification. which implies that the positive charges and positions of these lysine residues play an important role in a multipoint interaction of thaumatin I with human GCPRs. and experiment confirmed this predication as LTP1 had clear ice-binding activity and LTP2 did not (Doxey et al. Differences in the topology and surface electrostatic potential around the cleft are considered to determine the specificity of TLPs to their target proteins and ligands (Min et al. 2004). 2000).3-glucanase activity (MenuBouaouichea et al. Chemical modification found that five lysine residues (Lys78. 2006). Lys137 and Lys187) of thaumatin I contribute markedly to its sweetness (Kaneko and Kitabatake 2001). are separate and spread over a broad surface region around the cleft. 2006). Kalpana et al. Despite a high identity between two LTP amino acid sequences.Plant Cell Rep (2010) 29:419–436 431 cleft of other TLPs with b-1.Transgenic tobacco plants with the thaumatin gene exhibited delayed disease symptoms caused by Pythium aphanidermatum and Rhizoctonia solani. an algorithm revealed a clear ice-binding face on one (LTP1). For example. This His22 is situated on a loop of the protein surface which distinguishes TLX1 from other. The challenge remains to identify and explain fully what the functions of these proteins are. 2007). transgenic strawberry plants expressing the thaumatin II gene showed a significantly higher level of resistance to gray mold (Botrytis cinerea) (Schestibratov and Dolgov 2005). transgenic tobacco plants showed enhanced tolerance to necrotization caused by the pathogen Alternaria alternata (Velazhahan and Muthukrishnan 2003). but not on the other (LTP2). it appears that transgenic expression of thaumatin could be useful for modifying the taste of a variety of foods and plants bearing edible fruits (Masudaa and Kitabatake 2006). but did not disrupt the fold of the icebinding face (Middleton et al. Fagoaga et al. A field test of creeping bentgrass lines with rice D34 demonstrated that the transgenic plants displayed an improved resistance to the causal agent of dollar spot (Sclerotinia 123 . It is common that different members may have been coopted to additional functions during evolution of a multiple PR gene family. The rice TLP (D34) showed a similar effect in transgenic tobacco: at high D34 expression levels. thaumatin has a basic cleft structure. non-inhibiting TLPs. On the other hand. 2009). Transgenic cucumber (Cucumis sativus L.

Singh NK. chitinase. 2003). Bansal KC (2001) Overexpression of osmotin induces proline accumulation and confers tolerance to osmotic stress in transgenic tobacco. Robertus JD (1996) The crystal structure of the antifungal protein zeamatin. Van Leuven F. Similar to related TLPs. osmatin has a structurally homologous lectin-like b-barrel domain (Min et al. suggesting that expression of AtTLP1 in the roots is not required for systemic expression of ISR in the ´ leaves (Leon-Kloosterziel et al. Trick HN. Niedziela A. Transgenic cucumbers with an accumulation of the thaumatin II protein and a sweeter phenotype did not exhibit tolerance to the pathogenic fungus Pseudoperonospora cubensis (Maria et al. Niemirowicz-Szczytt K (2003) Modification of tomato taste in transgenic plants carrying a thaumatin gene from Thaumatococcus daniellii Benth. chitin-binding ¨ protein from Streptomyces tendae Tu901 that interferes with growth polarity. suggesting disease resistance may be the effect of multiple defence-responsive proteins (Lorito et al. D’Urzo MP. Hasegawa PM. In a field test with inoculation of S. Baier D. a member of the thaumatin-like. 2005). Plant Breed 122:347–351 Batalia MA. 2005). but no improvement against brown patch (Rhizoctonia solani) under greenhouse conditions (Fu et al. 2005). Gill BS. Babu V. overlapped with that of adiponectin. Zhou T. Tegoni M. Muthukrishnan S (2003) Greenhouse and field testing of transgenic wheat plants stably expressing genes for thaumatin-like protein. Unfortunately. Monzingo AF. Ernst S. and glucanase did not show resistance to Fusarium-caused scab under field conditions (Anand et al. 2004). Schwarz H (1999) Characterization of a novel. Van Damme EJM. Narasimhan ML. Bockus WW. This emerging knowledge indicates great potential for the application of TLP genes in gene engineering technologies for crop improvement and for use in medicine. May ´ GD. J Plant Biochem Biotechnol 10:31–37 Bartoszewski G. Cambillau C. the synergistic interaction of TLPs with other defence-related proteins. Plant Sci 118:11–23 Anand A. 2005). Niu X. chitinase and glucanase against Fusarium graminearum. arteriosclerosis. These results indicate the application of TLP genes to engineering disease resistance in plants needs to consider many factors. Conclusion Additional research and data generated in the past decade have led to a better understanding of the TLP family in variety of organisms. Herrera AF.432 Plant Cell Rep (2010) 29:419–436 homoeocarpa). heart disease. Zhu JK. PR-5 protein family. Horr I. 2007). transgenic wheat plants stably expressing genes for TLP. Plants transformed with TLPs do not always show enhanced pathogen resistance. and the complexity of ecosystems. Allergy 59:479–481 123 . 2003). information on fundamental aspects of the TLP family remains quite limited. could be used as a surrogate for adiponectin in the development of new therapeutic agents for the treatment of a wide variety of mammalian disorders involved in the adiponectin receptormediated pathway. Acknowledgments This work is supported in part by the Canadian Forest Service and the CFS-Genome R&D Initiative. such as the specialization of TLPs on pathogen range. However. 1996). FEBS Lett 572:3–7 Breiteneder H (2004) Thaumatin-like proteins—a new family of pollen and fruit allergens. and others (Narasimhan et al. cerevisiae and activates AMP kinase in C2C12 myocytes via a receptor that has homology to mammalian receptors for the hormone adiponectin (Narasimhan et al. Planta 211:791–799 Barthakur S. This b-barrel domain in the TLP structure. 2003). Berger J. especially the biological functions of TLPs relative to disease resistance and adaptation to stressful environments. 2002). Dermastia M (2003) The comparative analysis of osmotins and osmotin-like PR-5 proteins. Menu-Bouaouiche L. Pelosi P (2004) Plant stress proteins of the thaumatin-like family discovered in animals. AtTLP1 knockout mutant plants showed normal levels of rhizobacteriainduced systemic resistance (ISR) against the bacterial leaf pathogen. Rouge P (2000) Purification and structural analysis of an abundant thaumatin-like protein from ripe banana fruit. Angeli S. Nat Struct Biol 3:19–23 ¨ Bormann C. antifungal. Reuveni M. Bressen RA (1996) Antifungal activity of tobacco osmotin has specificity and involves plasma membrane permeabilization. Liu D. Such disorders include diabetes. Roberts W. Jung G. Osmatin induces apoptosis in the yeast S. Application of TLPs as mammalian therapeutic agents The antifungal capability of various TLPs suggests the existence of a signaling or recognition role mediated by their specific interaction with targets at the fungal plasma membrane (Veronese et al. J Bacteriol 181:7421–7429 Brandazza A. Peumans WJ. Plant Biol 5:116–124 Barre A. J Exp Bot 54:1101–1111 ˇ Anzlovar S. particularly on the precise mechanisms of protein regulation and function. This means that challenging fundamental and applied studies need to be conducted to characterize TLP receptors and their interacting components in the many life processes affected by this complex protein superfamily. Coexpression of TLP and chitinase in the progenies of a transgenic rice line revealed an enhanced resistance to the sheath blight pathogen (Rhizoctonia solani) as compared to transgenic lines expressing the individual genes (Maruthasalam et al. Raps C. homoeocarpa. some transgenic lines of creeping bentgrass with AtPR5K overexpression also showed delays in disease development of 29–45 days as relative to the control plants (Guo et al. References Abad LR. Szwacka M.

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