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Characterization of a lily anther-specific gene encoding cytoskeleton-binding glycoproteins and overexpression of the gene causes severe inhibition of pollen tube growth

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Abstract

This work characterizes an anther/pollen-specific gene that encodes potential intermediate filament (IF)-binding glycoproteins in lily (Lilium longiflorum Thunb. cv. Snow Queen) anthers during the development and pollen germination. LLP13 is a single gene that encodes a polypeptide of 807 amino acids, and a calculated molecular mass of 91 kDa. The protein contains a predicted transmembrane domain at the N-terminus and a conserved domain of unknown function (DUF)593 at the C-terminal half of the polypeptide. Sequence analysis revealed that LLP13 shares significant identity (37–41 %) with two intermediate filament antigen-binding proteins, representing a unique subgroup of DUF593 domain proteins from known rice and Arabidopsis species. The expression of LLP13 gene is anther-specific, and the transcript accumulates only at the stage of pollen maturation. Both premature drying and abscisic acid (ABA) treatment of developing pollen indicated that LLP13 was not induced by desiccation and ABA, but by other developmental cues. Antiserum was raised against the overexpressed LLP13C fragment of the protein in Escherichia coli and affinity-purified antibodies were prepared. Immunoblot analyses revealed that the LLP13 protein was a heterogeneous, anther-specific glycoprotein that accumulated only at the stage of pollen maturation. The protein is not heat-soluble. The level of LLP13 protein remained for 24 h during germination in vitro. Overexpression of LLP13-GFP or GFP-LLP13 in lily pollen tubes caused severe inhibition of tube elongation. The LLP13 protein codistributed with mTalin in growing tubes, suggesting that it apparently decorates actin cytoskeleton and is likely a cytoskeleton-binding protein that binds with IFs that potentially exist in pollen tubes.

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Abbreviations

ABA:

Abscisic acid

ConA:

Concanavalin A

GFP:

Green fluorescent protein

HRP:

Horseradish peroxidase

IF:

Intermediate filament

IFABP:

Intermediate filament antigen-binding protein

mTalin:

Mouse talin

NTA:

Nitrilotriacetic acid

RFP:

Red fluorescent protein

References

  • Allwood EG, Anthony RG, Smertenko AP, Reichelt S, Drobak BK, Doonan JH, Weeds AG, Hussey PJ (2002) Regulation of the pollen-specific actin-depolymerizing factor LlADF1. Plant Cell 14:2915–2927

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Altschul SF, Madden TL, Schafer AA, Zhang J, Zhang Z, Miller W, Lipman DJ (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25:3389–3402

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Becker JD, Boavida LC, Carneiro J, Haury M, Feijo JA (2003) Transcriptional profiling of Arabidopsis tissues reveals the unique characteristics of the pollen transcriptome. Plant Physiol 133:713–725

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Chen CY, Wong EI, Vidali L, Estavillo A, Hepler PK, Wu HM, Cheung AY (2002) The regulation of actin organization by actin depolymerizing factor in elongating pollen tubes. Plant Cell 14:2175–2190

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Cheung AY, Wu HM (2008) Structural and signaling networks for the polar cell growth machinery in pollen tubes. Annu Rev Plant Biol 59:547–572

    Article  CAS  PubMed  Google Scholar 

  • Cheung AY, Niroomand S, Zou Y, Wu HM (2010) A transmembrane formin nucleates subapical actin assembly and controls tip-focused growth in pollen tubes. Proc Natl Acad Sci USA 107:16390–16395

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Chiang JY, Balic N, Hsu SW, Yang CY, Ko CW, Hsu YF, Swoboda I, Wang CS (2006) A pollen-specific polygalacturonase from lily is related to major grass pollen allergens. Plant Physiol Biochem 44:743–751

    CAS  PubMed  Google Scholar 

  • Clegg JC (1982) Glycoprotein detection in nitrocellulose transfers of electrophoretically separated protein mixtures using concanavalin A and peroxidase: application to arenavirus and flavivirus proteins. Anal Biochem 127:389–394

    Article  CAS  PubMed  Google Scholar 

  • Dawson PJ, Hulme JS, Lloyd CW (1985) Monoclonal antibody to intermediate filament antigen crossreacts with higher plant cells. J Cell Biol 100:1793–1798

    Article  CAS  PubMed  Google Scholar 

  • Dickinson DB (1978) Influence of borate and pentaerythritol concentrations on germination and tube growth of Lilium longiflorum pollen. J Am Soc Horti Sci 103:413–416

    CAS  Google Scholar 

  • Fu Y, Wu G, Yang Z (2001) Rop GTPase-dependent dynamics of tip-localized F-actin controls tip growth in pollen tubes. J Cell Biol 152:1019–1032

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Gardiner J, Overall R, Marc J (2011) Putative Arabidopsis homologues of metazoan coiled-coil cytoskeletal proteins. Cell Biol Int 35:767–774

    CAS  PubMed  Google Scholar 

  • Guan Y, Guo J, Li H, Yang Z (2013) Signaling in pollen tube growth: crosstalk, feedback, and missing links. Mol Plant 6:1053–1064

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Habgreaves AJ, Dawson PJ, Butcher GW, Larkins A, Goodbody KC, Lloyd CW (1989) A monoclonal antibody raised against cytoplasmic fibrillar bundles from carrot cells, and its cross-reaction with animal intermediate filaments. J Cell Sci 92:371–378

    Google Scholar 

  • Holding DR, Otegui MS, Li B, Meeley RB, Dam T, Hunter BG, Jung R, Larkins BA (2007) The maize floury1 gene encodes a novel endoplasmic reticulum protein involved in zein protein body formation. Plant Cell 19:2569–2582

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Honys D, Twell D (2003) Comparative analysis of the Arabidopsis pollen transcriptome. Plant Physiol 132:640–652

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Hsu YF, Wang CS, Raja R (2007) Gene expression pattern at desiccation in the anther of Lilium longiflorum. Planta 226:311–322

    Article  CAS  PubMed  Google Scholar 

  • Hsu SW, Cheng CL, Tzen TJ, Wang CS (2010) Rop GTPase and its target cdc42/Rac-interactive-binding motif-containing protein genes respond to desiccation during pollen maturation. Plant Cell Physiol 51:1197–1209

    CAS  PubMed  Google Scholar 

  • Huang JC, Lin SM, Wang CS (2000) Characterization of a pollen-specific and desiccation-associated transcript in Lilium longiflorum during development and stress. Plant Cell Physiol 41:477–485

    CAS  PubMed  Google Scholar 

  • Huang S, Jin L, Du J, Li H, Zhao Q, Ou G, Ao G, Yuan M (2007) SB401, a pollen-specific protein from Solanum berthaultii, binds to and bundles microtubules and F-actin. Plant J 51:406–418

    Article  CAS  PubMed  Google Scholar 

  • Hussey PJ, Allwood EG, Smertenko AP (2002) Actin-binding proteins in the Arabidopsis genome database: properties of functionally distinct plant actin-depolymerizing factors/cofilins. Philos Trans R Soc Lond B Biol Sci 357:791–798

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Jia H, Li J, Zhu J, Fan T, Qian D, Zhou Y, Wang J, Ren H, Xiang Y, An L (2013) Arabidopsis CROLIN1, a novel plant actin binding protein, functions in cross-linking and stabilizing actin filaments. J Biol Chem 288:32277–32288

    Article  CAS  PubMed  Google Scholar 

  • Kost B, Spielhofer P, Chua NH (1998) A GFP-mouse talin fusion protein labels plant actin filaments in vivo and visualizes the actin cytoskeleton in growing pollen tubes. Plant J 16:393–401

    Article  CAS  PubMed  Google Scholar 

  • Ma BY, Qian D, Nan Q, Tan C, An LZ, Xiang Y (2012) Arabidopsis V-ATPase B subunits are involved in actin cytoskeleton remodeling via binding to, bundling and stabilizing F-actin. J Biol Chem 287:19008–19017

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Maliga P, Klessig DF, Cashmore AR, Gruissem W, Varner JE (1995) Methods in plant molecular biology. A laboratory course manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor

    Google Scholar 

  • McCormick S (1993) Male gametophyte development. Plant Cell 5:1265–1275

    Article  PubMed Central  PubMed  Google Scholar 

  • McCormick S (2004) Control of male gametophyte development. Plant Cell 16(Suppl):S142–S153

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Meagher RB, Fechheimer M (2003) The Arabidopsis cytoskeletal genome. Arabidopsis Book 2:e0096

    Article  PubMed Central  PubMed  Google Scholar 

  • Palanivelu R, Preuss D (2000) Pollen tube targeting and axon guidance: parallels in tip growth mechanisms. Trends Cell Biol 10:517–524

    Article  CAS  PubMed  Google Scholar 

  • Pina C, Pinto F, Feijo JA, Becker JD (2005) Gene family analysis of the Arabidopsis pollen transcriptome reveals biological implications for cell growth, division control, and gene expression regulation. Plant Physiol 138:744–756

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Ross JHE, Hutchings A, Butcher GW, Lane EB, Lloyd CW (1991) The intermediate filament-related system of higher plant cells shares an epitope with cytokeratin 8. J Cell Sci 99:91–98

    CAS  Google Scholar 

  • Roychoudhury A, Paul S, Basu S (2013) Cross-talk between abscisic acid-dependent and abscisic acid-independent pathways during abiotic stress. Plant Cell Rep 32:985–1006

    Article  CAS  PubMed  Google Scholar 

  • Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual, 2nd edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor

    Google Scholar 

  • Smith DE, Fisher PA (1984) Identification, developmental regulation, and response to heat shock of two antigenetically related forms of a major nuclear envelope protein in Drosophila embryos. Application of an improved method for affinity purification of antibodies using polypeptides immobilized on nitrocellulose blots. J Cell Biol 99:20–28

    Article  CAS  PubMed  Google Scholar 

  • Strelkov SV, Herrmann H, Geisler N, Wedig T, Zimbelmann R, Aebi U, Burkhard P (2002) Conserved segments 1A and 2B of the intermediate filament dimer: their atomic structures and role in filament assembly. EMBO J 21:1255–1266

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Sumner JB, Gralén N, Eriksson-Quensel IB (1938) The molecular weights of concanavalin A and concanavalin B. J Biol Chem 125:45–48

    CAS  Google Scholar 

  • Twell D, Yamaguchi J, Wing R, Ushiba J, McCormick S (1991) Promoter analysis of genes that are coordinately expressed during pollen development reveals pollen-specific enhancer sequences and shared regulatory elements. Genes Dev 5:496–507

    Article  CAS  PubMed  Google Scholar 

  • Wang CS, Walling LL, Eckard KJ, Lord EM (1992a) Patterns of protein accumulation in developing anthers of Lilium longiflorum correlate with histological events. Am J Bot 79:118–127

    Article  CAS  Google Scholar 

  • Wang CS, Walling LL, Eckard KJ, Lord EM (1992b) Immunological characterization of a tapetal protein in developing anthers of Lilium longiflorum. Plant Physiol 99:822–829

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Wang CS, Wu TD, Chung CKW, Lord EM (1996) Two classes of pollen-specific, heat-stable proteins in Lilium longiflorum. Physiol Plant 97:643–650

    Article  CAS  Google Scholar 

  • Wang HJ, Wan AR, Jauh GY (2008) An actin-binding protein, LlLIM1, mediates calcium and hydrogen regulation of actin dynamics in pollen tubes. Plant Physiol 147:1619–1636

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Whippo CW, Khurana P, Davis PA, DeBlasio SL, DeSloover D, Staiger CJ, Hangarter RP (2011) THRUMIN1 is a light-regulated actin-bundling protein involved in chloroplast motility. Curr Biol 21:59–64

    Article  CAS  PubMed  Google Scholar 

  • Wu YJ, Yan J, Zhang RH, Qu XL, Ren SL, Chen NZ, Huang SJ (2010) Arabidopsis FIMBRIN5, an actin bundling factor, is required for pollen germination and pollen tube growth. Plant Cell 22:3745–3763

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Yang CY, Chen YC, Jauh GY, Wang CS (2005) A lily ASR protein involves abscisic acid signaling and confers drought and salt resistance in Arabidopsis. Plant Physiol 139:836–846

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Zhao Y, Zhao SS, Mao TL, Qu XL, Cao WH, Zhang L, Zhang W, He L, Li SD, Ren SL, Zhao JF, Zhu GL, Huang SJ, Ye KQ, Yuan M, Guo Y (2011) The plant-specific actin binding protein SCAB1 stabilizes actin filaments and regulates stomatal movement in Arabidopsis. Plant Cell 23:2314–2330

    Article  PubMed Central  PubMed  Google Scholar 

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Acknowledgments

This research was supported by National Science Council of the Republic of China (Taiwan) under a Grant NSC101-2311-B-005-004-MY3 to Co-Shine Wang.

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Authors and Affiliations

  1. Graduate Institute of Biotechnology, National Chung Hsing University, Taichung, 40227, Taiwan

    Bing-Jyun Wang, Yi-Feng Hsu, Yun-Chu Chen & Co-Shine Wang

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  1. Bing-Jyun Wang
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  2. Yi-Feng Hsu
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Correspondence to Co-Shine Wang.

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Wang, BJ., Hsu, YF., Chen, YC. et al. Characterization of a lily anther-specific gene encoding cytoskeleton-binding glycoproteins and overexpression of the gene causes severe inhibition of pollen tube growth. Planta 240, 525–537 (2014). https://doi.org/10.1007/s00425-014-2099-x

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  • DOI: https://doi.org/10.1007/s00425-014-2099-x

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