鹅p21基因克隆、启动子分析及转录调控研究

doi:10.16431/j.cnki.1671-7236.2022.02.001

Abstract

Abstract: 【Objective】 The aim of this study was to analyze the structure and promoter activity of p21 gene in goose, and investigate the transcriptional regulation mechanism.【Method】 The Taizhou goose was taken as the research object, the sequence characteristics of the full-length and 5′-flanking region of p21 gene were obtained using homologous amplification, RACE and biological analysis.The dual luciferase reporter vectors with 6 different deletion fragments of the promoter were constructed, its luciferase activities were analyzed, and the core promoter region of p21 gene was identified.Site-directed mutagenesis of the transcription factor binding sites MyoD (+25 to +36 bp) in the core promoter region was carried out, and the key transcription factors of p21 gene was preliminary identified in C2C12 cell line.【Result】 The full cDNA sequence of p21 gene in goose was 1 943 bp, containing 453 bp coding regions (CDS) which encoded 151 amino acids.p21 protein sequence had highly conserved CDI family binding sites.Phylogenetic tree analysis showed that p21 gene in goose was closely related to Anas platyrhynchos, and had a particularly strong evolutionary relationship with Gallus gallus and Meleagris gallopavo.The 5′-flanking region of p21 gene contained promoter elements, the core promoter was found to be located at —35 to +37 bp, which played a positive regulatory effect.Combined with site-directed mutation demonstrated that MyoD was the key transcription factor of p21 gene in goose.【Conclusion】 The complete cDNA sequence and promoter region of p21 gene in goose were obtained, and MyoD was the key transcription factor.The results provided a theoretical basis for exploring molecular regulation mechanism of p21 gene in embryonic muscle development of goose.

Key words: goose; p21 gene; cloning; promoter activity; transcriptional regulatory element

CLC Number:

CHEN Zhe, YANG Pengxia, LEI Mingming, CHEN Jiabin, YAN Leyan. Cloning, Promoter Analysis and Transcriptional Regulation of p21 Gene in Goose[J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(2): 405-415.

References

[1] CHINZEI N, HAYASHI S, UEHA T, et al. p21 deficiency delays regeneration of skeletal muscular tissue[J]. PLoS One, 2015, 10(5): e0125765.
[2] 陈晓霞. CDKN1A、CDKN1B基因在蛋鸡等级前卵泡组织中的时空表达模式研究[J]. 中国家禽, 2019, 41(4): 12-16.
CHEN X X. Temporal and spatial expression patterns of CDKN1A and CDKN1B genes in pre-hierarchical follicle of laying hens[J]. China Poultry, 2019, 41(4): 12-16. (in Chinese)
[3] 郑海英, 黄玥萌, 杨春艳, 等. 水牛p21基因在卵母细胞体外成熟过程中的表达及其真核表达载体构建[J]. 动物医学进展, 2018, 39(10): 18-24.
ZHENG H Y, HUANG Y M, YANG C Y, et al. Expression of p21 gene in water buffalo oocytes during in vitro maturation and construction of its eukaryotic expression vector[J]. Progress in Veterinary Science, 2018, 39(10): 18-24. (in Chinese)
[4] 宋锦, 姬牧远, 吴西军, 等. p21基因在野生型斑马鱼胚胎发育过程中的表达[J]. 贵州医科大学学报, 2016, 41(7): 765-769.
SONG J, JI M Y, WU X J, et al. The expression of p21 in development of wild type zebrafish embryo[J]. Journal of Guizhou Medical University, 2016, 41(7): 765-769. (in Chinese)
[5] 李鹂, 黄衡宇, 谢永芳. p21基因在小鼠胚胎发育过程中的表达[J]. 吉首大学学报(自然科学版), 2006, 27(2): 80-83.
LI L, HUANG H Y, XIE Y F. Study of gene expression of p21 with in situ hybridization technique[J]. Journal of Jishou University (Natural Sciences Edition), 2006, 27(2): 80-83. (in Chinese)
[6] ISHIDO M, KAMI K, MASUHARA M. In vivo expression patterns of MyoD, p21 and Rb proteins in myonuclei and satellite cells of denervated rat skeletal muscle[J]. American Journal of Physiology-Cell Physiology, 2004, 287(2): C484-C493.
[7] 孙荣距, 夏照帆, 杨宗城, 等. E-box元件在p21基因表达调控中作用的研究[J]. 感染、炎症、修复, 2008, 9(2): 90-93.
SUN R J, XIA Z F, YANG Z C, et al. Study on effect of the element of E-box in regulating the expression of p21 gene[J]. Infection, Inflammation, Repair, 2008, 9(2): 90-93. (in Chinese)
[8] GARTEL A L, TYNER A L. Transcriptional regulation of the p21 (WAF1/CIP1) gene[J]. Experimental Cell Research, 1999, 246(2): 280-289.
[9] JIN J J, LV W, XIA P, et al. Long noncoding RNA SYISL regulates myogenesis by interacting with polycomb repressive complex 2[J]. Proceedings of the National Academy of Sciences of the United States of America, 2018, 115(42): E9802-E9811.
[10] 郭倩倩, 王大涛, 褚文辉, 等. p21基因与哺乳动物器官再生相关性研究[J]. 中国畜牧兽医, 2014, 41(5): 171-176.
GUO Q Q, WANG D T, CHU W H, et al. Research progress of relationship between p21 gene and mammalian regeneration[J]. China Animal Husbandry & Veterinary Medicine, 2014, 41(5): 171-176. (in Chinese)
[11] KARIMIAN A, AHMADI Y, YOUSEFI B. Multiple functions of p21 in cell cycle, apoptosis and transcriptional regulation after DNA damage[J]. DNA Repair, 2016, 42: 63-71.
[12] 王小斌. KLF7基因调控肌肉卫星细胞静息的分子机制研究[D]. 杨凌: 西北农林科技大学, 2016.
WANG X B. Molecular mechanism of KLF7 gene regulation on muscle satellite cell quiescence[D]. Yangling: Northwest A&F University, 2016. (in Chinese)
[13] KUROKAWA M, SATO F, ARAMAKI S, et al. Monitor of the myostatin antocrine action during differentiation of embryonic chicken myoblasts into myotubes: Effect of IGF-1[J]. Molecular and Cellular Biochemistry, 2009, 331(1-2): 193-199.
[14] PICARD B, LEFAUCHEUR L, BERRI C, et al. Muscle fiber ontogenesis in farm animal species[J]. Reproduction Nutrition Development, 2002, 42(5): 415-431.
[15] 刘宏祥, 徐文娟, 朱春红, 等. 鸭胚胎发育中后期胸肌发育阻滞的RNA-seq分析[J]. 中国农业科学, 2018, 51(22): 4373-4386.
LIU H X, XU W J, ZHU C H, et al. RNA-seq analysis on development arrest of duck pectoralis muscle during semi-late embryonic period[J]. Scientia Agricultura Sinica, 2018, 51(22): 4373-4386. (in Chinese)
[16] O'NEILL J. Growth and differentiation during myogenesis in the chicken embryo[J]. Developmental Biology, 1987, 120(2): 465-540.
[17] 梁耀伟, 赵宗胜, 陈丹盈, 等. 鸡、鹌鹑及其杂交禽胚胎肌肉发育过程中MSTN和p21基因的表达规律[J]. 畜牧兽医学报, 2012, 43(6): 997-1002.
LIANG Y W, ZHAO Z S, CHEN D Y, et al. The expression profiles of MSTN and p21 during embryonic myogenesis in chicken, quail and hybrids of chicken-quail[J]. Acta Veterinaria et Zootechnica Sinica, 2012, 43(6): 997-1002. (in Chinese)
[18] 王婧, 李冰, 刘翠翠, 等. 启动子结构和功能研究进展[J]. 生物技术通报, 2014, 8(8): 40-45.
WANG J, LI B, LIU C C, et al. Advances of the studies on structure and function of promoter[J]. Biotechnology Bulletin, 2014, 8(8): 40-45. (in Chinese)
[19] HAELENS A, VERRIJDT G, SCHOENMAKERS E, et al. The first exon of human sc gene contains an androgen responsive unit and an interferon regulatory factor element[J]. Molecular and Cellular Endocrinology, 1999, 153(1): 91-102.
[20] GUO K, WANG J, ANDRES V, et al. MyoD-induced expression of p21 inhibits cyclin-dependent kinase activity upon myocyte terminal differentiation[J]. Molecular and Cellular Biology, 1995, 15(7): 3823-3829.
[21] LAST T J, BIRNBAUM M, VAN WIJNEN A J, et al. Repressor elements in the coding region of the human histone H4 gene interact with the transcription factor CDP/cu[J]. Gene, 1998, 221(2): 267-277.
[22] RAHMAN M, HIRABAYASHI Y, ISHⅡ T, et al. A repressor element in the 5'-untranslated region of human Pax5 exon 1A[J]. Gene, 2001, 263(1): 59-66.
[23] WARDLE F C. Master control: Transcriptional regulation of mammalian MyoD[J]. Journal of Muscle Research and Cell Motility, 2019, 40(2): 211-226.
[24] BERKES C A, TAPCCOTTS J. MyoD and the transcriptional control of myogenesis[J]. Seminars in Cell and Developmental Biology, 2005, 16(4): 585-595.
[25] CHOI J, COSTA M L, MERMELSTEIN C S, et al. MyoD converts primary dermal fibroblasts, chondroblasts, smooth muscle, and retinal pigmented epithelial cells into striated mononucleated myoblasts and multinucleated myotubes[J]. Proceedings of the National Academy of Sciences of the United States of America, 1990, 87(20): 7988-7992.
[26] HALEVY O, NOVITCH B G, SPICER D B, et al. Correlation of terminal cell cycle arrest of skeletal muscle with induction of p21 by MyoD[J]. Science, 1995, 267(5200): 1018-1021.
[27] AGARAM N P, LAQUAGLIA M P, ALAGGIO R, et al. MyoD1-mutant spindle cell and sclerosing rhabdomyosarcoma: An aggressive subtype irrespective of age. A reappraisal for molecular classification and risk stratification[J]. Modern Pathology, 2019, 32(1): 27-36.
[28] BERGSTROM D A, TAPSCOTT S J. Molecular distinction between specification and differentiation in the myogenic basic helix-loop-helix transcription factor family[J]. Molecular and Cellular Biology, 2001, 21(7): 2404-2412.

Cloning, Promoter Analysis and Transcriptional Regulation of p21 Gene in Goose

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