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Journal of Integrative Agriculture  2014, Vol. 13 Issue (10): 2102-2112    DOI: 10.1016/S2095-3119(13)60643-7
Crop Genetics · Breeding · Germplasm Resources Advanced Online Publication | Current Issue | Archive | Adv Search |
Isolation and Expression Analysis of Two Genes Encoding Cinnamate 4-Hydroxylase from Cotton (Gossypium hirsutum)
 NI  Zhi-yong, LI  Bo, Neumann  M Peter, LÜ  Meng , FAN  Ling
1、Institute of Nuclear and Biological Technologies, Xinjiang Academy of Agricultural Sciences, Urumqi 830091, P.R.China
2、College of Agronomy, Xinjiang Agricultural University, Urumqi 830052, P.R.China
3、Plant Physiology Laboratory, Department of Environmental, Water and Agricultural Engineering, Faculty of Civil and Environmental
Engineering, Technion-Israel Institute of Technology, Haifa 32000, Israel
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摘要  Two genes (GhC4H1 and GhC4H2) that encode putative cotton cinnamate 4-hydroxylases that catalyze the second step in the phenylpropanoid pathway were isolated from developing cotton fibers. GhC4H1 and GhC4H2 each contain open reading frames of 1 518 base pairs (bp) in length and both encode proteins consisting of 505 amino acid residues. They are 90.89% identical to each other at the amino acid sequence level and belong to class I of plant C4Hs. GhC4H1 and GhC4H2 genomic DNA are 2 247 and 2 161 bp long, respectively, and contain two introns located at conserved positions relative to the coding sequence. GhC4H1 and GhC4H2 promoters were isolated and found to contain many cis-elements (boxes P, L and AC-I element) previously identified in the promoters of other phenylpropanoid pathway genes. Histochemical staining showed GUS expression driven by the GhC4H1 and GhC4H2 promoters in ovules and fibers tissues. GhC4H1 and GhC4H2 were also widely expressed in other cotton tissues. GhC4H2 expression reached its highest level during the elongation stage of fiber development, whereas GhC4H1 expression increased during the secondary wall development period in cotton fibers. Our results contribute to a better understanding of the biochemical role of GhC4H1 and GhC4H2 in cotton fiber development.

Abstract  Two genes (GhC4H1 and GhC4H2) that encode putative cotton cinnamate 4-hydroxylases that catalyze the second step in the phenylpropanoid pathway were isolated from developing cotton fibers. GhC4H1 and GhC4H2 each contain open reading frames of 1 518 base pairs (bp) in length and both encode proteins consisting of 505 amino acid residues. They are 90.89% identical to each other at the amino acid sequence level and belong to class I of plant C4Hs. GhC4H1 and GhC4H2 genomic DNA are 2 247 and 2 161 bp long, respectively, and contain two introns located at conserved positions relative to the coding sequence. GhC4H1 and GhC4H2 promoters were isolated and found to contain many cis-elements (boxes P, L and AC-I element) previously identified in the promoters of other phenylpropanoid pathway genes. Histochemical staining showed GUS expression driven by the GhC4H1 and GhC4H2 promoters in ovules and fibers tissues. GhC4H1 and GhC4H2 were also widely expressed in other cotton tissues. GhC4H2 expression reached its highest level during the elongation stage of fiber development, whereas GhC4H1 expression increased during the secondary wall development period in cotton fibers. Our results contribute to a better understanding of the biochemical role of GhC4H1 and GhC4H2 in cotton fiber development.
Keywords:  cinnamate 4-hydroxylase       Gossypium hirsutum       promoter analysis       phenylpropanoid pathway  
Received: 26 June 2013   Accepted:
Fund: 

This work was funded by the National Natural Science Foundation of China (31060173), the Joint Funds of the National Natural Science Foundation of China (U1178305) and the High-Tech R&D Program of Xinjiang, China (201111116).

Corresponding Authors:  FAN Ling, Tel/Fax: +86-991-4527003, E-mail: fanling@xaas.ac.cn     E-mail:  fanling@xaas.ac.cn
About author:  NI Zhi-yong, E-mail: nizhiyong@126.com;

Cite this article: 

NI Zhi-yong, LI Bo, Neumann M Peter, Lü Meng , FAN Ling. 2014. Isolation and Expression Analysis of Two Genes Encoding Cinnamate 4-Hydroxylase from Cotton (Gossypium hirsutum). Journal of Integrative Agriculture, 13(10): 2102-2112.

Al-Ghazi Y, Bourot S, Arioli T, Dennis E S, Llewellyn DJ. 2009. Transcript profiling during fiber developmentidentifies pathways in secondary metabolism and cell wallstructure that may contribute to cotton fiber quality. Plantand Cell Physiology, 50, 1364-1381

Basra A S, Malik C P. 1984. Development of the cottonfiber. International Review of Cytology-A Survey of CellBiology, 89, 65-113

Bell-Lelong D A, Cusumano J C, Meyer K, ChappleC. 1997. Cinnamate-4-hydroxylase expression inArabidopsis Regulation in response to development andthe environment. Plant Physiology, 113, 729-738

Betz C, McCollum T G, Mayer R T. 2001. Differentialexpression of two cinnamate 4-hydroxylase genes in‘Valencia’ orange (Citrus sinensis Osbeck). PlantMolecular Biology, 46, 741-748

Chapple C. 1998. Molecular-genetic analysis of plantcytochrome P450-dependent monooxygenases. AnnualReview of Plant Physiology and Plant Molecular Biology,49, 311-343

Chen A H, Chai Y R, Li J N, Chen L. 2007. Molecularcloning of two genes encoding cinnamate 4-hydroxylase(C4H) from oilseed rape (Brassica napus). Journal ofBiochemistry and Molecular Biology, 40, 247-260

Dixon R A, Paiva N L. 1995. Stress-induced phenylpropanoidmetabolism. The Plant Cell, 7, 1085-1097

Durst F, OKeefe D P. 1995. Plant cytochromes P450: Anoverview. Drug Metabolism and Drug Interaction, 12,171-187

Fan L, Linker R, Gepstein S, Tanimoto E, Yamamoto R,Neumann P M. 2006. Progressive inhibition by waterdeficit of cell wall extensibility and growth along theelongation zone of maize roots is related to increased ligninmetabolism and progressive stelar accumulation of wallphenolics. Plant Physiology, 140, 603-612

Fan L, Shi W J, Hu W R, Hao X Y, Wang D M, Yuan H,Yan H Y. 2009. Molecular and biochemical evidence forphenylpropanoid synthesis and presence of wall-linkedphenolics in cotton fibers. Journal of Integrative PlantBiology, 51, 626-637

Frank M R, Deyneka J M, Schuler M A. 1996. Cloning ofwound-induced cytochrome P450 monooxygenasesexpressed in pea. Plant Physiology, 110, 1035-1046

Gou J Y, Wang L J, Chen S P, Hu W L, Chen X Y. 2007. Geneexpression and metabolite profiles of cotton fiber duringcell elongation and secondary cell wall syntheisis. CellResearch, 17, 422-434

Hovav R, Udall J, Hovav E, Rapp R, Flagel L, Wendel J. 2008.A majority of cotton genes are expressed in single-celledfiber. Planta, 227, 319-329

Huang B, Duan Y, Yi B, Sun L, Lu B, Yu X, Sun H, Zhang H,Chen W. 2008. Characterization and expression profilingof cinnamate 4-hydroxylase gene from Salvia miltiorrhizain rosmarinic acid biosynthesis pathway. Russian Journalof Plant Physiology, 55, 390-399

Jefferson R A, Kavanagh T A, Bevan M W. 1987. GUS fusions:b-glucuronidase as a sensitive and versatile gene fusionmarker in higher plants. The EMBO Journal, 6, 3901-3907

Jin H, Cominelli E, Bailey P, Parr A, Mehrtens F, Jones J,Tonelli C, Weisshaar B, Martin C. 2000. Transcriptionalrepression by AtMYB4 controls production of UVprotectingsunscreens in Arabidopsis. The EMBO Journal,19, 6150-6161

John M E, Keller G. 1996. Metabolic pathway engineering incotton: biosynthesis of polyhydroxybutyrate in fiber cells.Proceedings of the National Academy of Sciences of theUnited States of America, 93, 12768-12773

Kim H J, Williams M Y, Triplett B A. 2002. A novel expression assay system for fiber-specific promoters in developingcotton fibers. Plant Molecular Biology Reporter, 20, 7-18

Koopman E, Logemann E, Hahlbrock K. 1999. Regulation andfunctional expression of cinnamate 4-hydroxylase fromparsley. Plant Physiology, 119, 49-55

Liu S, Hu Y, Wang X, Han L, Song S, Cheng H, Lin Z.2009. Isolation and characterization of a gene encodingcinnamate 4-hydroxylase from Parthenocissus henryana.Molecular Biology Reports, 36, 1605-1610

Lois R, Dietrich A, Hahlbrock K, Schulz W. 1989. Aphenylalanine ammonia-lyase gene from parsley:Structure, regulation and identification of elicitor andlight responsive cis-acting elements. The EMBO Journal,8, 1641-1648

Lu S, Zhou Y, Li L, Chiang V L. 2006. Distinct roles ofcinnamate 4-hydroxylase genes in Populus. Plant and CellPhysiology, 47, 905-914

Luo P, Wang G D, Chen X Y. 2001. Isolation and expressionanalysis of two cDNAs encoding C4H homologues fromGossypium arboreum. Acta Botanica Sinica, 43, 77-81

Mizutani M, Ohta D, Sato R. 1997. Isolation of a cDNA anda genomic clone encoding cinnamate 4-hydroxylase fromArabidopsis and its expression manner in planta. PlantPhysiology, 11, 755-763

Mizutani M, Ward E, DiMaio J, Ohta D, Ryals J, Sato R. 1993.Molecular cloning and sequencing of a cDNA encodingmung bean cytochrome P450 (P450C4H) possessingcinnamate 4-hydroxylase activity. Biochemical andBiophysical Research Communications, 190, 875-880

Nedelkina S, Jupe S C, Blee K A, Schalk M, Daniele W R,Bolwell G P. 1999. Novel characteristics and regulationof a divergent cinnamate 4-hydroxylase (CYP73A15)from French bean: Engineering expression in yeast. PlantMolecular Biology, 39, 1079-1090

Nelson D R, Schuler M A, Paquette S M, Werck-ReichhartD, Bak S. 2004. Comparative genomics of rice andArabidopsis. Analysis of 727 cytochrome P450 genes andpseudogenes from a monocot and a dicot. Plant Physiology,135, 756-772

Ro D K, Douglas C J. 2004. Reconstitution of the entrypoint of plant phenylpropanoid metabolism in yeast(Saccharomyces cerevisiae): Implications for control ofmetabolic flux into the phenylpropanoid pathway. Journalof Biological Chemistry, 279, 2600-2607

Ro D K, Mah N, Ellis B E, Douglas C J. 2001. Functionalcharacterization and subcellular localization of poplar(Populus trichocarpa×Populus deltoides) cinnamate4-hydroxylase. Plant Physiology, 126, 317-329

Schoch G A, Attias R, le Ret M, Werck-Reichhart D. 2003.Key substrate recognition residues in the active site ofa plant cytochrome P450, CYP73A1: Homology modelguided site-directed mutagenesis. The FEBS Journal, 270,3684-3695

Shi Y H, Zhu S W, Mao X Z, Feng J X, Qin Y M, ZhangL, Cheng J, Wei L P, Wang Z Y, Zhu Y X. 2006.Transcriptome profiling, molecular biological, andphysiological studies reveal a major role for ethylene incotton fiber cell elongation. The Plant Cell, 18, 651-664

Tan J F, Tu L L, Deng F L, Hu H Y, Nie Y C, Zhang X L. 2013.A genetic and metabolic analysis revealed that cotton fibercell development was retarded by flavonoid naringenin.Plant Physiology, 162, 86-95

Teutsch H G, Hasenfratz M P, Lesot A, Stoltz C, Garnier J M,Jeltsch J M, Durst F, Werck-Reichhart D. 1993. Isolationand sequence of a cDNA encoding the Jerusalem artichokecinnamate 4-hydroxylase, a major plant cytochromeP450 involved in the general phenylpropanoid pathway.Proceedings of the National Academy of Sciences of theUnited States of America, 90, 4102-4106

Urban P, Werck-Reichhart D, Teutsch H G, Durst F, RegnierS, Kazmaier M, Pompon D. 1994. Characterization ofrecombinant plant cinnamate 4-hydroxylase produced inyeast. Kinetic and spectral properties of the major plantP450 of the phenylpropanoid pathway. European Journalof Biochemistry, 222, 843-850

Wan C Y, Wilkins T A. 1994. A modified hot borate methodsignificantly enhances the yield of high-quality RNA fromcotton (Gossypium hirsutum L.). Analytical Biochemistry,223, 7-12

Weisshaar B, Jenkins G I. 1998. Phenylpropanoid biosynthesisand its regulation. Current Opinion in Plant Biology, 1,251-257

Yamazaki S, Sato K, Suhara K, Sakaguchi M, Mihara K,Omura T. 1993. Importance of the proline-rich regionfollowing signal anchor sequence in the formation ofcorrect conformation of microsomal cytochrome P450s.Journal of Biochemistry, 114, 652-657

Yang D H, Chung B Y, Kim J S, Kim J H, Yun P Y, Lee YK, Lim Y P, Lee M C. 2005. cDNA cloning and sequenceanalysis of the rice cinnamate-4-hydroxylase gene, acytochrome P450-dependent monooxygenase involvedin the general phenylpropanoid pathway Journal of PlantBiology, 48, 311-318

Zhong R, Ye Z H. 2009. Transcriptional regulation of ligninbiosynthesis. Plant Signaling Behavior, 4, 1028-1034

Zhou J, Lee C, Zhong R, Ye Z H. 2009. MYB58 and MYB63are transcriptional activators of the lignin biosyntheticpathway during secondary cell wall formation inArabidopsis. The Plant Cell, 21, 248-266
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