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Molecular Genetics and Genomics

, Volume 293, Issue 5, pp 1191–1204 | Cite as

Ectopic expression of GhCOBL9A, a cotton glycosyl-phosphatidyl inositol-anchored protein encoding gene, promotes cell elongation, thickening and increased plant biomass in transgenic Arabidopsis

  • Erli Niu
  • Shuai Fang
  • Xiaoguang Shang
  • Wangzhen GuoEmail author
Original Article

Abstract

Cellulose is a major component of plant cell walls and is necessary for plant morphogenesis and biomass. COBL (COBRA-Like) proteins have been shown to be key regulators in the orientation of cell expansion and cellulose crystallinity status. To clarify the role of a cotton COBL gene, GhCOBL9A, we conducted the ectopic expression and functional analysis in Arabidopsis. Previous study showed that GhCOBL9A was preferentially expressed during secondary cell wall biosynthesis in cotton fibers, and showed a significant co-expression pattern with cellulose synthase genes. Here, we detected that overexpression of GhCOBL9A induced the up-regulation of genes related to cellulose synthesis and enhanced the cellulose deposition. As a result, GhCOBL9A transgenic plants displayed increased hypocotyl and root lengths in early development, and cell wall thickening at the SCW stage. Notably, overexpression of GhCOBL9A led to an erect, robust-stature phenotype and brought higher biomass in mature plants. In addition, overexpression of GhCOBL9A in Arabidopsis AtCOBL4 mutants, a paralogous gene of GhCOBL9A, also led to a stronger growth potential, but the Atcobl4 mutant phenotype could not be rescued, implying the functional divergence of GhCOBL9A and AtCOBL4 paralogs. Taken together, these results suggest that overexpression of GhCOBL9A contributes to plant cell elongation and thickening, and increased biomass, which provides references for further utilizing GhCOBL9A to improve yield and quality traits in cotton and other species.

Keywords

GhCOBL9A Cell elongation Cellulose deposition Biomass Functional divergence 

Abbreviations

PCW

Primary cell wall

SCW

Secondary cell wall

Susy

Sucrose synthase

KOR

Korrigan

CTL

Chitinase-like

CesAs

Cellulose synthases

COBL

COBRA-Like

GPI

Glycosyl-phosphatidyl inositol

CBM

Carbohydrate-binding module

CSCs

Cellulose synthesizing complexes

CaMV

Cauliflower mosaic virus

DPA

Days post anthesis

qRT-PCR

Quantitative real-time PCR

PIECE

Plant intron exon comparison and evolution database

CDD

Conserved domain database

ORF

Open reading frame

MS

Murashige and Skoog

BLAST

Basic local alignment search tool

ph

Phloem

xy

Xylem

if

Interfascicular fiber

pi

Pith cell

Notes

Acknowledgements

This program was financially supported in part by National Natural Science Foundation of China (31701472), Natural Science Foundation in Jiangsu Province (BK20160712), and Jiangsu Collaborative Innovation Center for Modern Crop Production (No. 10).

Compliance with ethical standards

Conflict of interest

The authors declared they had no conflict of interest.

Ethical approval

The experiments in this manuscript complied with the current laws of the country in which they were performed.

Supplementary material

438_2018_1452_MOESM1_ESM.tiff (145 kb)
Fig. S1 Identification of GhCOBL9A transgenic lines in Arabidopsis. Detection on GhCOBL9A overexpression transgenic lines in DNA (a) and transcription (b) levels. WT, Arabidopsis Columbia-0 (Col-0); OE1 to OE10, overexpression GhCOBL9A transgenic lines in WT plants. The expression level of AtUbq5 (NM_116090.3) was used as the internal control and the relative expression level was calculated using the 2-△CT method (Livak and Schmittgen 2001) (TIFF 144 KB)
438_2018_1452_MOESM2_ESM.tiff (95 kb)
Fig. S2 Number of different types of branches in GhCOBL9A transgenic plants. Different Arabidopsis branches were referred to the descriptions as Sugimoto et al. (2014) (TIFF 95 KB)
438_2018_1452_MOESM3_ESM.tiff (140 kb)
Fig. S3 Identification of GhCOBL9A transgenic lines in Atcobl4 mutant. Detection on GhCOBL9A overexpression transgenic lines in DNA (a) and transcription (b) levels. Atcobl4, T-DNA insertion mutant of AtCOBL4 (IRX6; N431557); RM1 to RM12, overexpression GhCOBL9A transgenic lines in Atcobl4 mutant. The expression level of AtUbq5 (NM_116090.3) was used as the internal control and the relative expression level was calculated using the 2-△CT method (Livak and Schmittgen 2001) (TIFF 139 KB)

References

  1. Adams KL, Wendel JF (2005) Polyploidy and genome evolution in plants. Curr Opin Plant Biol 8:135–141CrossRefPubMedGoogle Scholar
  2. Amor Y, Haigler CH, Johnson S, Wainscott M, Delmer DP (1995) A membrane-associated form of sucrose synthase and its potential role in synthesis of cellulose and callose in plants. Proc Natl Acad Sci USA 92:9353–9357CrossRefPubMedGoogle Scholar
  3. Benfey PN, Linstead PJ, Roberts K, Schiefelbein JW, Hauser MT, Aeschabacher RA (1993) Root development in Arabidopsis: four mutants with dramatically altered root morphogenesis. Development 119:57–70PubMedGoogle Scholar
  4. Ben-Tov D, Abraham Y, Stav S, Thompson K, Loraine A, Elbaum R, de Souza A, Pauly M, Kieber JJ, Harpaz-Saad S (2015) COBRA-LIKE2, a member of the glycosylphosphatidylinositol-anchored COBRA-LIKE family, plays a role in cellulose deposition in Arabidopsis seed coat mucilage secretory cells. Plant Physiol 167:711–724CrossRefPubMedPubMedCentralGoogle Scholar
  5. Brady SM, Song S, Dhugga KS, Rafalski JA, Benfey PN (2007) Combining expression and comparative evolutionary analysis. The COBRA gene family. Plant Physiol 143:172–187CrossRefPubMedPubMedCentralGoogle Scholar
  6. Brown DM, Zeef LAH, Ellis J, Goodacre R, Turner SR (2005) Identification of novel genes in Arabidopsis involved in secondary cell wall formation using expression profiling and reverse genetics. Plant Cell 17:2281–2295CrossRefPubMedPubMedCentralGoogle Scholar
  7. Ching A, Dhugga KS, Appenzeller L, Meeley R, Bourret TM, Howard RJ, Rafalski A (2006) Brittle stalk 2 encodes a putative glycosylphosphatidylinositol-anchored protein that affects mechanical strength of maize tissues by altering the composition and structure of secondary cell walls. Planta 224:1174–1184CrossRefPubMedGoogle Scholar
  8. Clough SJ, Bent AF (1998) Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. Plant J 16:735–743CrossRefPubMedGoogle Scholar
  9. Dai XX, You CJ, Wang L, Chen GX, Zhang QF, Wu CY (2009) Molecular characterization, expression pattern, and function analysis of the OsBC1L family in rice. Plant Mol Biol 71:469–481CrossRefPubMedGoogle Scholar
  10. Dai XX, You CJ, Chen GX, Li XH, Zhang QF, Wu CY (2011) OsBC1L4 encodes a COBRA-like protein that affects cellulose synthesis in rice. Plant Mol Biol 75:333–345CrossRefPubMedGoogle Scholar
  11. Guerriero G, Fugelstad J, Bulone V (2010) What do we really know about cellulose biosynthesis in higher plants? J Integr Plant Biol 52:161–175CrossRefPubMedGoogle Scholar
  12. Gutierrez L, Mauriat M, Guénin S, Pelloux J, Lefebvre JF, Louvet R, Rusterucci C, Moritz T, Guerineau F, Bellini C, Van Wuytswinkel O (2008) The lack of a systematic validation of reference genes: a serious pitfall undervalued in reverse transcription-polymerase chain reaction (RT-PCR) analysis in plants. Plant Biotechnol J 6:609–618CrossRefPubMedGoogle Scholar
  13. Haigler CH, Betancur L, Stiff MR, Tuttle JR (2012) Cotton fiber: a powerful single-cell model for cell wall and cellulose research. Front Plant Sci 3:104CrossRefPubMedPubMedCentralGoogle Scholar
  14. Hajdukiewicz P, Svab Z, Maliga P (1994) The small, versatile pPZP family of Agrobacterium binary vectors for plant transformation. Plant Mol Biol 25:989–994CrossRefPubMedGoogle Scholar
  15. Hauser MT, Morikami A, Benfey (1995) Conditional root expansion mutants of Arabidopsis. Development 121:1237–1252PubMedPubMedCentralGoogle Scholar
  16. Higgins DG, Thompson JD, Gibson TJ (1996) Using CLUSTAL for multiple sequence alignments. Methods Enzymol 266:383–402CrossRefPubMedGoogle Scholar
  17. Hochholdinger F, Wen TJ, Zimmermann R, Chimot-Marolle P, da Costa e Silva O, Bruce W, Lamkey KR, Wienand U, Schnable PS (2008) The maize (Zea mays L.) roothairless 3 gene encodes a putative GPI-anchored, monocot-specific, COBRA-like protein that significantly affects grain yield. Plant J 54:888–898CrossRefPubMedPubMedCentralGoogle Scholar
  18. Houston K, Tucker MR, Chowdhury J, Shirley N, Little A (2016) The plant cell wall: a complex and dynamic structure as revealed by the responses of genes under stress conditions. Front Plant Sci 7:984CrossRefPubMedPubMedCentralGoogle Scholar
  19. Hughes J, McCully ME (1975) The use of an optical brightener in the study of plant structure. Stain Technol 50:319–329CrossRefPubMedGoogle Scholar
  20. Hussey SG, Mizrachi E, Creux NM, Myburg AA (2013) Navigating the transcriptional roadmap regulating plant secondary cell wall deposition. Front Plant Sci 4:325CrossRefPubMedPubMedCentralGoogle Scholar
  21. Jiang JX, Zhang TZ (2003) Extraction of total RNA in cotton tissues with CTAB-acidic phenolic method. Cotton Sci 15:166–167Google Scholar
  22. Kim HJ, Triplett BA (2001) Cotton fiber growth in planta and in vitro: models for plant cell elongation and cell wall biogenesis. Plant Physiol 127:1361–1366CrossRefPubMedPubMedCentralGoogle Scholar
  23. Kumar M, Turner S (2015) Plant cellulose synthesis: CESA proteins crossing kingdoms. Phytochemistry 112:91–99CrossRefPubMedGoogle Scholar
  24. Kumar M, Campbell L, Turner S (2016) Secondary cell walls: biosynthesis and manipulation. J Exp Bot 67:515–531CrossRefPubMedGoogle Scholar
  25. Kumar M, Atanassov I, Turner S (2017) Functional analysis of cellulose synthase (CESA) protein class specificity. Plant Physiol 173:970–983CrossRefPubMedGoogle Scholar
  26. Li YH, Qian Q, Zhou YH, Yan MX, Sun L, Zhang M, Fu ZM, Wang YH, Han B, Pang XM, Chen MS, Li JY (2003) BRITTLE CULM1, which encodes a COBRA-like protein, affects the mechanical properties of rice plants. Plant Cell 15:2020–2031CrossRefPubMedPubMedCentralGoogle Scholar
  27. Li SD, Bashline L, Lei L, Gu Y (2014) Cellulose synthesis and its regulation. Arabidopsis Book 12:e0169CrossRefPubMedPubMedCentralGoogle Scholar
  28. Liu L, Shang-Guan K, Zhang B, Liu X, Yan M, Zhang L, Shi Y, Zhang M, Qian Q, Li J, Zhou Y (2013) Brittle Culm1, a COBRA-like protein, functions in cellulose assembly through binding cellulose microfibrils. PLoS Genet 9:e1003704CrossRefPubMedPubMedCentralGoogle Scholar
  29. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2–△△CT method. Methods 25:402–408CrossRefPubMedPubMedCentralGoogle Scholar
  30. Marchler-Bauer A, Bo Y, Han L, He J, Lanczycki CJ, Lu S, Chitsaz F, Derbyshire MK, Geer RC, Gonzales NR, Gwadz M, Hurwitz DI, Lu F, Marchler GH, Song JS, Thanki N, Wang Z, Yamashita RA, Zhang D, Zheng C, Geer LY, Bryant SH (2017) CDD/SPARCLE: functional classification of proteins via subfamily domain architectures. Nucleic Acids Res 45:D200–D203CrossRefPubMedGoogle Scholar
  31. Mitsuda N, Seki M, Shinozaki K, Ohme-Takagi M (2005) The NAC transcription factors NST1 and NST2 of Arabidopsis regulate secondary wall thickenings and are required for anther dehiscence. Plant Cell 17:2993–3006CrossRefPubMedPubMedCentralGoogle Scholar
  32. Nicol F, His I, Jauneau A, Vernhettes S, Canut H, Hofte H (1998) A plasma membrane-bound putative endo-1,4-beta-D-glucanase is required for normal wall assembly and cell elongation in Arabidopsis. EMBO J 17:5563–5576CrossRefPubMedPubMedCentralGoogle Scholar
  33. Niu E, Shang X, Cheng C, Bao J, Zeng Y, Cai C, Du X, Guo W (2015) Comprehensive analysis of the COBRA-Like (COBL) gene family in Gossypium identifies two COBLs potentially associated with fiber quality. PLoS One 10:e0145725CrossRefPubMedPubMedCentralGoogle Scholar
  34. O’Brien TP, Feder N, McCully ME (1964) Polychromatic staining of plant cell walls by toluidine blue O. Protoplasma 59:368–373CrossRefGoogle Scholar
  35. Paterson AH, Brubaker CL, Wendel JF (1993) A rapid method for extraction of cotton (Gossypium spp.) genomic DNA suitable for RFLP or PCR analysis. Plant Mol Biol Rep 11:122–127CrossRefGoogle Scholar
  36. Provenzano M, Mocellin S (2007) Complementary techniques: validation of gene expression data by quantitative real time PCR. Adv Exp Med Biol 593:66–73CrossRefPubMedGoogle Scholar
  37. Richmond TA, Somerville CR (2000) The cellulose synthase superfamily. Plant Physiol 124:495–498CrossRefPubMedPubMedCentralGoogle Scholar
  38. Roudier F, Schindelman G, DeSalle R, Benfey PN (2002) The COBRA family of putative GPI-anchored proteins in Arabidopsis. a new fellowship in expansion. Plant Physiol 130:538–548CrossRefPubMedPubMedCentralGoogle Scholar
  39. Roudier F, Fernandez AG, Fujita M, Himmelspach R, Borner GH, Schindelman G, Song S, Baskin TI, Dupree P, Wasteneys GO, Benfey PN (2005) COBRA, an Arabidopsis extracellular glycosyl-phosphatidyl inositol-anchored protein, specifically controls highly anisotropic expansion through its involvement in cellulose microfibrils orientation. Plant Cell 17:1749–1763CrossRefPubMedPubMedCentralGoogle Scholar
  40. Sánchez-Rodríguez C, Bauer S, Hématy K, Saxe F, Ibáñez AB, Vodermaier V, Konlechner C, Sampathkumar A, Rüggeberg M, Aichinger E, Neumetzler L, Burgert I, Somerville C, Hauser MT, Persson S (2012) Chitinase-like1/POM-POM1 and its homolog CTL2 are glucan-interacting proteins important for cellulose biosynthesis in Arabidopsis. Plant Cell 24:589–607CrossRefPubMedPubMedCentralGoogle Scholar
  41. Scheible WR, Pauly M (2004) Glycosyltransferases and cell wall biosynthesis: novel players and insights. Curr Opin Plant Biol 7:285–295CrossRefPubMedGoogle Scholar
  42. Schindelman G, Morikami A, Jung J, Baskin TI, Carpita NC, Derbyshire P, McCann MC, Benfey PN (2001) COBRA encodes a putative GPI-anchored protein, which is polarly localized and necessary for oriented cell expansion in Arabidopsis. Genes Dev 5:1115–1127CrossRefGoogle Scholar
  43. Shang X, Chai Q, Zhang Q, Jiang J, Zhang T, Guo W, Ruan Y (2015) Down-regulation of the cotton endo-1,4-β-glucanase gene KOR1 disrupts endosperm cellularization, delays embryo development, and reduces early seedling vigour. J Exp Bot 66:3071–3083CrossRefPubMedPubMedCentralGoogle Scholar
  44. Somerville C (2006) Cellulose synthesis in higher plants. Annu Rev Cell Dev Biol 22:53–78CrossRefPubMedGoogle Scholar
  45. Somerville C, Bauer S, Brininstool G, Facette M, Hamann T, Milne J, Osborne E, Paredez A, Persson S, Raab T, Vorwerk S, Youngs H (2004) Toward a systems approach to understanding plant cell walls. Science 306:2206–2211CrossRefPubMedGoogle Scholar
  46. Sugimoto H, Kondo S, Tanaka T, Imamura C, Muramoto N, Hattori E, Ogawa K, Mitsukawa N, Ohto C (2014) Overexpression of a novel Arabidopsis PP2C isoform, AtPP2CF1, enhances plant biomass production by increasing inflorescence stem growth. J Exp Bot 65:5385–5400CrossRefPubMedPubMedCentralGoogle Scholar
  47. Taylor-Teeples M, Lin L, de Lucas M, Turco G, Toal TW, Gaudinier A, Young NF, Trabucco GM, Veling MT, Lamothe R, Handakumbura PP, Xiong G, Wang C, Corwin J, Tsoukalas A, Zhang L, Ware D, Pauly M, Kliebenstein DJ, Dehesh K, Tagkopoulos I, Breton G, Pruneda-Paz JL, Ahnert SE, Kay SA, Hazen SP, Brady SM (2015) An Arabidopsis gene regulatory network for secondary cell wall synthesis. Nature 517:571–575CrossRefPubMedGoogle Scholar
  48. Updegraff DM (1969) Semimicro determination of cellulose in biological materials. Anal Biochem 32:420–424CrossRefPubMedGoogle Scholar
  49. Wang Y, Xu L, Thilmony R, You FM, Gu YQ, Coleman-Derr D (2017) PIECE 2.0: an update for the plant gene structure comparison and evolution database. Nucleic Acids Res 45:1015–1020CrossRefPubMedGoogle Scholar
  50. Wendel JF (2000) Genome evolution in polyploids. Plant Mol Biol 42:225–249CrossRefPubMedGoogle Scholar
  51. Zhang HB, Li YN, Wang BH, Chee PW (2008) Recent advances in cotton genomics. Int J Plant Genom 2008:742304Google Scholar
  52. Zhong R, Ye ZH (2015) Secondary cell walls: biosynthesis, patterned deposition and transcriptional regulation. Plant Cell Physiol 56:195–214CrossRefPubMedGoogle Scholar
  53. Zhong R, Richardson EA, Ye ZH (2007) The MYB46 transcription factor is a direct target of SND1 and regulates secondary wall biosynthesis in Arabidopsis. Plant Cell 19:2776–2792CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.State Key Laboratory of Crop Genetics and Germplasm Enhancement, Hybrid Cotton R & D Engineering Research Center, Ministry of EducationNanjing Agricultural UniversityNanjingChina

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