Plant Cell, Tissue and Organ Culture (PCTOC)

, Volume 110, Issue 2, pp 261–273 | Cite as

Overexpression of Arabidopsis cyclin D2;1 in cotton results in leaf curling and other plant architectural modifications

  • Guanze Liu
  • Shuangxia Jin
  • Xuyan Liu
  • Jiafu Tan
  • Xiyan Yang
  • Xianlong Zhang
Original Paper


The D-class cyclin genes play key roles in controlling the cell cycle in plant development. To evaluate the effects of D-type cyclins on plant architecture, ectopic expression of the Arabidopsis cyclinD2;1 (Arath;CYCD2;1) gene driven by the CaMV35S promoter was investigated in cotton via Agrobacterium-mediated transformation. Northern blot showed the cyclinD2;1 gene was highly expressed in transgenic cotton plants. Phenotype investigation showed that overexpression of Arath;CYCD2;1 led to obvious leaf architecture change: the leaf epidermis of transgenic plants consist of more small cells compared to the wild-type. Mesophyll cells in the inner layers of Arath;CYCD2;1 plants were organized more loosely than those in the inner layers of wild-type plants. Moreover, transgenic plants had darker leaves, more chlorophyll, and a higher rate of photosynthesis than wild-type plants in the field. Tissue cultures indicated that the overexpression of Arath;CYCD2;1 promoted callus formation in the absence of exogenous auxin, but inhibited cell differentiation. The qRT-PCR revealed that several cell cycle–associated genes, particularly the transcript levels of GhRBR and GhCYCD3;1 were regulated by the Arath;CYCD2;1 insertion. The results implied that the cotton plant architecture or cell culture characters could be regulated by ectopic expression of the Arabidopsis cyclinD2;1 gene.


D-type cyclins Arath;CYCD2;1 Leaf rolling Plant architecture Transgenic cotton 



Cauliflower mosaic virus


Cyclin-dependent kinase


Inhibitor of cyclin-dependent kinase


Retinoblastoma-related protein


N-1-naphthylphthalamic acid


Minichromosomal maintenance



The authors thank Dr. Y. M. Zhou for his generosity in providing the CycD2 cDNA vector and Dr. J. L. Yao for her help with histological observation. This work was supported by the National Natural Science Foundation of China (No. 30810103911).


  1. Arnon DI (1949) Cooper enzymes isolated chloroplasts polyphenol oxidase in Beta vulgaris. Plant Physiol 24:1–15PubMedCrossRefGoogle Scholar
  2. Beemster GTS, Mironov V, Inze D (2005) Tuning the cell-cycle engine for improved plant performance. Curr Opin Biotechnol 16:142–146PubMedCrossRefGoogle Scholar
  3. Borghi L, Gutzat R, Futterer J, Laizet YH, Hennig L, Gruissem W (2010) Arabidopsis RETINOBLASTOMA-RELATED is required for stem cell maintenance, cell differentiation, and lateral organ production. Plant Cell 22:1792–1811PubMedCrossRefGoogle Scholar
  4. Brendel V, Kleffe J, Carle Urioste JC, Walbot V (1998) Prediction of splice sites in plant pre-mRNA from sequence properties. J Mol Biol 276:85–104PubMedCrossRefGoogle Scholar
  5. Cockcroft CE, den Boer BGW, Healy JM, Murray JAH (2000) Cyclin D control of growth rate in plants. Nature 405:575–579PubMedCrossRefGoogle Scholar
  6. De Veylder L, Beeckman T, Inze D (2007) The ins and outs of the plant cell cycle. Nat Rev Mol Cell Biol 8:655–665PubMedCrossRefGoogle Scholar
  7. Desvoyes B, Ramirez Parra E, Xie Q, Chua NH, Gutierrez C (2006) Cell type-specific role of the retinoblastoma/E2F pathway during Arabidopsis leaf development. Plant Physiol 140:67–80PubMedCrossRefGoogle Scholar
  8. Dewitte W, Riou Khamlichi C, Scofield S, Healy JM, Jacqmard A, Kilby NJ, Murray JAH (2003) Altered cell cycle distribution, hyperplasia, and inhibited differentiation in Arabidopsis caused by the D-type cyclin CYCD3. Plant Cell 15:79–92PubMedCrossRefGoogle Scholar
  9. Ebel C, Mariconti L, Gruissem W (2004) Plant retinoblastoma homologues control nuclear proliferation in the female gametophyte. Nature 429:776–780PubMedCrossRefGoogle Scholar
  10. Falster DS, Westoby M (2003) Leaf size and angle vary widely across species: what consequences for light interception? New Phytol 158:509–525CrossRefGoogle Scholar
  11. Francis D (2007) The plant cell cycle—15 years on. New Phytol 174:261–278PubMedCrossRefGoogle Scholar
  12. Hu YX, Bao F, Li JY (2000) Promotive effect of brassinosteroids on cell division involves a distinct CycD3-induction pathway in Arabidopsis. Plant J 24:693–701PubMedCrossRefGoogle Scholar
  13. Huntley R, Healy S, Freeman D, Lavender P, de Jager S, Greenwood J, Makker J, Walker E, Jackman M, Xie Q, Bannister AJ, Kouzarides T, Gutiérrez C, Doonan JH, Murray JAH (1998) The maize retinoblastoma protein homologue ZmRb-1 is regulated during leaf development and displays conserved interactions with G1/S regulators and plant cyclin D (CycD) proteins. Plant Mol Biol 37:155–169PubMedCrossRefGoogle Scholar
  14. Inze D, De Veylder L (2006) Cell cycle regulation in plant development. Annu Rev Genet 40:77–105PubMedCrossRefGoogle Scholar
  15. Jin SX, Zhang XL, Liang SG, Nie YC, Guo XP, Huang C (2005) Factors affecting transformation efficiency of embryogenic callus of upland cotton (Gossypium hirsutum) with Agrobacterium tumefaciens. Plant Cell, Tissue Organ Cult 81:229–237CrossRefGoogle Scholar
  16. Jin SX, Zhang XL, Nie YC, Guo XP, Liang SG, Zhu HG (2006) Identification of a novel elite genotype for in vitro culture and genetic transformation of cotton. Biol Plantarum 50:519–524CrossRefGoogle Scholar
  17. Meijer M, Murray JAH (2001) Cell cycle controls and the development of plant form. Curr Opin Plant Biol 4:44–49PubMedCrossRefGoogle Scholar
  18. Menges M, Samland AK, Planchais S, Murray JAH (2006) The D-type cyclin CYCD3;1 is limiting for the G1-to-S-phase transition in Arabidopsis. Plant Cell 18:893–906PubMedCrossRefGoogle Scholar
  19. Menges M, Pavesi G, Morandini P, Bogre L, Murray JAH (2007) Genomic organization and evolutionary conservation of plant D-type cyclins. Plant Physiol 145:1558–1576PubMedCrossRefGoogle Scholar
  20. Montero-Cortés M, Rodríguez-Paredes F, Burgeff C, Pérez-Nuñez T, Córdova I, Oropeza C, Verdeil JL, Sáenz L (2010) Characterisation of a cyclin-dependent kinase (CDKA) gene expressed during somatic embryogenesis of coconut palm. Plant Cell, Tissue Organ Cult 102:251–258CrossRefGoogle Scholar
  21. Montero-Cortés M, Cordova I, Verdeil J-L, Hocher V, Pech y Ake A, Sandoval A, Oropeza C, Saenz L (2011) GA3 induces expression of E2F-like genes and CDKA during in vitro germination of zygotic embryos of Cocos nucifera (L). Plant Cell, Tissue Organ Cult 107:461–470CrossRefGoogle Scholar
  22. Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plantarum 15:473–497CrossRefGoogle Scholar
  23. Nakagami H, Kawamura K, Sugisaka K, Sekine M, Shinmyo A (2002) Phosphorylation of retinoblastoma-related protein by the cyclin D/cyclin-dependent kinase complex is activated at the G1/S-phase transition in tobacco. Plant Cell 14:1847–1857PubMedCrossRefGoogle Scholar
  24. Oakenfull EA, Riou Khamlichi C, Murray JAH (2002) Plant D-type cyclins and the control of G1 progression. Philos T Roy Soc B 357:749–760CrossRefGoogle Scholar
  25. Oh SJ, Kim SJ, Kim Y, Park SH, Ha SH, Kim JK (2008) Arabidopsis cyclin D2 expressed in rice forms a functional cyclin-dependent kinase complex that enhances seedling growth. Plant Biotechnol Rep 2:227–231CrossRefGoogle Scholar
  26. Park JA, Ahn JW, Kim YK, Kim SJ, Kim JK, Kim WT, Pai HS (2005) Retinoblastoma protein regulates cell proliferation, differentiation, and endoreduplication in plants. Plant J 42:153–163PubMedCrossRefGoogle Scholar
  27. Paul S, Dam A, Bhattacharyya A, Bandyopadhyay T (2011) An efficient regeneration system via direct and indirect somatic embryogenesis for the medicinal tree Murraya koenigii. Plant Cell, Tissue Organ Cult 105:271–283CrossRefGoogle Scholar
  28. Qi R, John PCL (2007) Expression of genomic AtCYCD2;1 in Arabidopsis induces cell division at smaller cell sizes: Implications for the control of plant growth. Plant Physiol 144:1587–1597PubMedCrossRefGoogle Scholar
  29. Riou-Khamlichi C, Huntley R, Jacqmard A, Murray JAH (1999) Cytokinin activation of Arabidopsis cell division through a D-type cyclin. Science 283:1541–1544PubMedCrossRefGoogle Scholar
  30. Riou-Khamlichi C, Menges M, Healy JM, Murray JAH (2000) Sugar control of the plant cell cycle: differential regulation of Arabidopsis D-type cyclin gene expression. Mol Cell Biol 20:4513–4521PubMedCrossRefGoogle Scholar
  31. Sanz L, Dewitte W, Forzani C, Patell F, Nieuwland J, Wen B, Quelhas P, De Jager S, Titmus C, Campilho A, Ren H, Estelle M, Wang H, Murray JAH (2011) The Arabidopsis D-type cyclin CYCD2;1 and the inhibitor ICK2/KRP2 modulate auxin-induced lateral root formation. Plant Cell 23:641–666PubMedCrossRefGoogle Scholar
  32. Shen LL, Chen Y, Su XH, Zhang SG, Pan HX, Huang MR (2012) Two FT orthologs from Populus simonii Carrière induce early flowering in Arabidopsis and poplar trees. Plant Cell, Tissue Organ Cult 108:371–379CrossRefGoogle Scholar
  33. Soni R, Carmichael JP, Shah ZH, Murray JAH (1995) A family of cyclin D homologs from plants differentially controlled by growth regulators and containing the conserved retinoblastoma protein interaction motif. Plant Cell 7:85–103PubMedGoogle Scholar
  34. Tsukaya H (2006) Mechanism of leaf-shape determination. Annu Rev Plant Biol 57:477–496PubMedCrossRefGoogle Scholar
  35. Wang F, Huo SN, Guo J, Zhang XS (2006) Wheat D-type cyclin Triae;CYCD2;1 regulate development of transgenic Arabidopsis plants. Planta 224:1129–1140PubMedCrossRefGoogle Scholar
  36. Wyrzykowska J, Schorderet M, Pien S, Gruissem W, Fleming AJ (2006) Induction of differentiation in the shoot apical meristem by transient overexpression of a retinoblastoma-related protein. Plant Physiol 141:1338–1348PubMedCrossRefGoogle Scholar
  37. Zeng FC, Zhang XL, Zhu LF, Tu LL, Guo XP, Nie YC (2006) Isolation and characterization of genes associated to cotton somatic embryogenesis by suppression subtractive hybridization and macroarray. Plant Mol Biol 60:167–183PubMedCrossRefGoogle Scholar
  38. Zhang GH, Xu Q, Zhu XD, Qian Q, Xue HW (2009) SHALLOT-LIKE1 is a KANADI transcription factor that modulates rice leaf rolling by regulating leaf abaxial cell development. Plant Cell 21:719–735PubMedCrossRefGoogle Scholar
  39. Zhou YM, Fowke LC, Wang H (2002) Plant CDK inhibitors: studies of interactions with cell cycle regulators in the yeast two-hybrid system and functional comparisons in transgenic Arabidopsis plants. Plant Cell Rep 20:967–975CrossRefGoogle Scholar
  40. Zhou YM, Wang H, Gilmer S, Whitwill S, Fowke LC (2003) Effects of co-expressing the plant CDK inhibitor ICK1 and D-type cyclin genes on plant growth, cell size and ploidy in Arabidopsis thaliana. Planta 216:604–613PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2012

Authors and Affiliations

  • Guanze Liu
    • 1
  • Shuangxia Jin
    • 1
  • Xuyan Liu
    • 1
  • Jiafu Tan
    • 1
  • Xiyan Yang
    • 1
  • Xianlong Zhang
    • 1
  1. 1.National Key Laboratory of Crop Genetic ImprovementHuazhong Agricultural UniversityWuhanPeople’s Republic of China

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