Cloning,Bioinformatics Analysis and Expression Analysis of FTL Gene in Yak

doi:10.7668/hbnxb.20194239

Abstract

Abstract:

The aim of this study was to understand the structure and function of ferritin light chain(FTL)gene in yak and its expression pattern in eight tissues of different age groups.The coding sequence(CDS)of FTL gene was cloned from the cDNA of liver tissue of adult(36 months old)Meiren yak,and bioinformatics analysis was carried out.The relative expression of FTL gene in heart,liver,spleen,lung,kidney,longissimus dorsi,fat and testis was detected by Real-time fluorescence quantitative(qRT-PCR).The results showed that the CDS region of FTL gene of Meiren yak was 528 bp,encoding 175 amino acids,and had the closest relationship with wild yak and cattle(99.8%).FTL protein was a stable hydrophilic secretory protein without signal peptide and transmembrane structure.It was mainly distributed in the cytoplasm.The secondary structure of the protein was mainly α-helix,which mainly interacted with FTH1,SCARA5,NCOA4 and other proteins.The results of qRT-PCR showed that the expression levels of FTL gene in adult cattle were liver,kidney,testis,spleen,lung,fat,longissimus dorsi and heart,respectively.The expression levels of FTL gene in fat and longissimus dorsi tissues of 7-day-old calves were the highest,and the expression level of FTL gene in liver tissue of adult cattle was the highest.By comparing the two stages,it was found that the expression levels of fat,longissimus dorsi and heart tissues decreased with the increase of age,while the expression levels of lung,spleen,testis,kidney and liver tissues increased with age.

Key words: Meiren yak, FTL, Gene cloning, Biological analysis, Tissue expression

ZHANG Juanxiang, MA Xiaoyong, LU Jianwei, REN Qingji, LUO Shengwei, XIAO Guangfeng, WANG Youhe, MA Xiaoming, LA Yongfu, GUO Xian, CHU Min, YAN Ping, LIANG Chunnian. Cloning,Bioinformatics Analysis and Expression Analysis of FTL Gene in Yak[J]. Acta Agriculturae Boreali-Sinica, 2023, 38(6): 192-200. doi: 10.7668/hbnxb.20194239.

/ / Recommend / Download Citations

URL: http://www.hbnxb.net/EN/10.7668/hbnxb.20194239

http://www.hbnxb.net/EN/Y2023/V38/I6/192

Figures/Tables 16

References 31

[1]	窦承贤, 方敏全, 梁晋涛, 李桂银. 铁蛋白结构和功能的生物信息学分析[J]. 生物技术, 2023, 33(1):33-37,67.doi:10.16519/j.cnki.1004-311x.2023.01.0006.
	Dou C X, Fang M Q, Liang J T, Li G Y. Bioinformatics analysis of the structure and function of ferritin[J]. Biotechnolog, 2023, 33(1):33-37,67.
[2]	Finazzi D, Arosio P. Biology of ferritin in mammals:an update on iron storage,oxidative damage and neurodegeneration[J]. Archives of Toxicolog, 2014, 88(10):1787-1802.doi:10.1007/s00204-014-1329-0. URL
[3]	纪强. AFB1对原代小鼠腹腔巨噬细胞的毒性作用及铁稳态机制[D]. 哈尔滨: 东北农业大学, 2020.doi:10.27010/d.cnki.gdbnu.2020.000433.
	Ji Q. Toxic effect of AFB1 on primary mouse peritoneal macrophages and iron homeostasis mechanism[D]. Harbin: Northeast Agricultural University, 2020.
[4]	Zhang N, Yu X Q, Xie J X, Xu H M. New insights into the role of ferritin in iron homeostasis and neurodegenerative diseases[J]. Molecular Neurobiolog, 2021, 58(6):2812-2823.doi:10.1007/s12035-020-02277-7.
[5]	Segrestin B, Moreno-Navarrete J M, Seyssel K, Alligier M, Meugnier E, Nazare J A, Vidal H, Fernandez-Real J M, Laville M. Adipose tissue expansion by overfeeding healthy men alters iron gene expression[J]. The Journal of Clinical Endocrinology and Metabolism, 2019, 104(3):688-696.doi:10.1210/jc.2018-01169. URL
[6]	颜丽芬. 超顺磁氧化铁标记对大鼠脂肪干细胞及其心肌样分化细胞转铁蛋白受体和铁蛋白表达的影响[D]. 广州: 南方医科大学, 2012.doi:10.3969/j.issn.1000-9965.2012.04.013.
	Yan L F. Effect of superparamagnetic iron oxide labeling on transferrin receptor and ferritin expression in rat adipose-derived stem cells and their cardiomyocyte-like differentiated cells[D]. Guangzhou: Southern Medical University, 2012.
[7]	Sammarco M C, Ditch S, Banerjee A, Grabczyk E. Ferritin L and H subunits are differentially regulated on a post-transcriptional level[J]. The Journal of Biological Chemistr, 2008, 283(8):4578-4587.doi:10.1074/jbc.m703456200.
[8]	曾广. FTL在非小细胞肺癌中的表达及临床意义的研究[D]. 天津: 天津医科大学, 2021.doi:10.27366/d.cnki.gtyku.2020.000854.
	Zeng G. Study on the expression and clinical significance of FTL in non-small cell lung cancer[D]. Tianjin: Tianjin Medical University, 2021.
[9]	Folgueras A R, Freitas-Rodríguez S, Ramsay A J, Garabaya C, Rodríguez F, Velasco G, López-Otín C. Matriptase-2 deficiency protects from obesity by modulating iron homeostasis[J]. Nature Communications, 2018, 9:1350.doi:10.1038/s41467-018-03853-1. pmid: 29636509
[10]	Yan H F, Liu Z Y, Guan Z A, Guo C. Deferoxamine ameliorates adipocyte dysfunction by modulating iron metabolism in ob/ob mice[J]. Endocrine Connections, 2018, 7(4):604-616.doi:10.1530/ec-18-0054. URL
[11]	Wang Y F, Zhu S, Hodgkinson V, Prohaska J R, Weisman G A, Gitlin J D, Petris M J. Maternofetal and neonatal copper requirements revealed by enterocyte-specific deletion of the Menkes disease protein[J]. American Journal of Physiology Gastrointestinal and Liver Physiolog, 2012, 303(11):G1236-G1244.doi:10.1152/ajpgi.00339.2012.
[12]	杨晓斌, 何莉莉, 张扬, 周瑞锋, 后康丽, 张芸芸, 王月丽, 李青青, 石斌刚, 胡江. 牦牛乳铁蛋白基因多态性及其与乳品质性状的关联分析[J]. 中国畜牧兽医, 2021, 48(5):1655-1663.doi:10.16431/j.cnki.1671-7236.2021.05.016
	Yang X B, He L L, Zhang Y, Zhou R F, Hou K L, Zhang Y Y, Wang Y L, Li Q Q, Shi B G, Hu J. Polymorphisms of LF gene and their association with milk quality traits in yak[J]. China Animal Husbandry & Veterinary Medicine, 2021, 48(5):1655-1663.
[13]	南青卓玛, 何克磊. 合作市美仁牦牛提纯复壮现状及发展对策[J]. 中国畜禽种业, 2021, 17(5):92-93.doi:10.3969/j.issn.1673-4556.2021.05.054.
	Zhuoma N Q, He K L. Present situation and development countermeasures of purification and rejuvenation of Meiren Yak in Hezuo City[J]. The Chinese Livestock and Poultry Breeding, 2021, 17(5):92-93.
[14]	杨小林, 陈勇, 朱友军, 邹万瑞, 龙泽军, 李胜男, 刘英. 舍饲牦牛、犏牛和放牧牦牛屠宰性状、肉质及其营养成分比较[J]. 湖北农业科学, 2019, 58(24):170-174.doi:10.14088/j.cnki.issn0439-8114.2019.24.041.
	Yang X L, Chen Y, Zhu Y J, Zou W R, Long Z J, Li S N, Liu Y. Comparative analysis of carcass traits,quality and nutrients of muscle among house-feeding yaks,yak-cattles and yaks with grazing[J]. Hubei Agricultural Sciences, 2019, 58(24):170-174.
[15]	胡金丽, 庄蕾, 吴森. 牦牛肉质性状相关候选基因研究进展[J]. 中国畜牧杂志, 2021, 57(8):10-15.doi:10.19556/j.0258-7033.20200803-04.
	Hu J L, Zhuang L, Wu S. Research progress on candidate genes related to meat quality traits of yak[J]. Chinese Journal of Animal Husbandr, 2021, 57(8):10-15.
[16]	喻晓丹. 猪肝脏总RNA快速提取方法[J]. 湖北农业科学, 2012, 51(3):621-623.doi:10.3969/j.issn.0439-8114.2012.03.055.
	Yu X D. Fast extraction of total RNA from pig liver[J]. Hubei Agricultural Sciences, 2012, 51(3):621-623.
[17]	Qualmann B, Kessels M M. The role of protein arginine methylation as post-translational modification on actin cytoskeletal components in neuronal structure and function[J]. Cells, 2021, 10(5):1079.doi:10.3390/cells10051079. URL
[18]	吴双虎, 王鹏飞, 李宏, 晁娟娟, 王战航, 朱俊儒, 安小鹏. 关中奶山羊FGF2基因克隆、生物信息学分析及组织表达研究[J]. 中国畜牧兽医, 2022, 49(12):1454-1463.doi:10.16431/j.cnki.1671-7236.2022.12.005.
	Wu S H, Wang P F, Li H, Chao J J, Wang Z H, Zhu J R, An X P. Cloning,bioinformatics analysis and tissue expression study of FGF2 gene in Guanzhong dairy goats[J]. China Animal Husbandry & Veterinary Medicine, 2022, 49(12):1454-1463.
[19]	Muhoberac B B, Vidal R.Iron,ferritin,hereditary ferritinopathy,and neurodegeneration[J]. Frontiers in Neuroscience, 2019, 13:1195.doi:10.3389/fnins.2019.01195. pmid: 31920471
[20]	Sonnendecker C, Wei R, Kurze E, Wang J P, Oeser T, Zimmermann W. Efficient extracellular recombinant production and purification of a Bacillus cyclodextrin glucanotransferase in Escherichia coli[J]. Microbial Cell Factories, 2017, 16(1):87.doi:10.1186/s12934-017-0701-1. pmid: 28526016
[21]	Silva B, Faustino P. An overview of molecular basis of iron metabolism regulation and the associated pathologies[J]. Biochimica et Biophysica Acta (BBA)-Molecular Basis of Disease, 2015, 1852(7):1347-1359.doi:10.1016/j.bbadis.2015.03.011. pmid: 25843914
[22]	王丽, 李万宏, 李发弟, 李熙成, 乐祥鹏. FTH1基因多态性与湖羊和小尾寒羊产羔数的关联分析[J]. 基因组学与应用生物学, 2020, 39(4):1529-1534.doi:10.13417/j.gab.039.001529.
	Wang L, Li W H, Li F D, Li X C, Le X P. Association of polymorphism within FTH1 with litter size of Hu sheep and small-tailed Han sheep[J]. China Industrial Economics, 2020, 39(4):1529-1534.
[23]	Ferreira C, Bucchini D, Martin M E, Levi S, Arosio P, Grandchamp B, Beaumont C. Early embryonic lethality of H ferritin gene deletion in mice[J]. Journal of Biological Chemistr, 2000, 275(5):3021-3024.doi:10.1074/jbc.275.5.3021.
[24]	郭立春, 曹国栋, 罗茜晴, 林靓, 赵宇含, 曾佑成, 张艺馨, 程青虹. NCOA4介导的铁蛋白自噬参与LPS诱导的心肌细胞铁死亡[J]. 陆军军医大学学报, 2022, 44(23):2401-2408.doi:10.16016/j.2097-0927.202204140.
	Guo L C, Cao G D, Luo X Q, Lin L, Zhao Y H, Zeng Y C, Zhang Y X, Cheng Q H. NCOA4-mediated ferritin autophagy is involved in LPS-induced ferroptosis in cardiomyocytes[J]. Journal of Army Medical Universit, 2022, 44(23):2401-2408.
[25]	史子敏, 朱安义, 洪正东. SCARA5的研究进展[J]. 广东医学, 2018, 39(24):3724-3726.doi:10.3969/j.issn.1001-9448.2018.24.033.
	Shi Z M, Zhu A Y, Hong Z D. Research progress of SCARA5[J]. Guangdong Medical Journal, 2018, 39(24):3724-3726.
[26]	龚静, 曾献春. 山羊FTL基因克隆、序列分析和组织表达研究[J]. 黑龙江畜牧兽医, 2022(15):58-63,136,137.doi:10.13881/j.cnki.hljxmsy.2021.11.0230.
	Gong J, Zeng X C. Study on cloning,sequence analysis and tissue expression of FTL gene in goats[J]. Heilongjiang Animal Science and Veterinary Medicine, 2022(15):58-63,136,137.
[27]	Laghaei R, Kowallis W, Evans D G, Coalson R D. Calculation of iron transport through human H-chain ferritin[J]. The Journal of Physical Chemistry A, 2014, 118(35):7442-7453.doi:10.1021/jp500198u. URL
[28]	Alfrey C P, Lynch E C, Whitley C E. Characteristics of ferritin isolated from human marrow,spleen,liver,and reticulocytes[J]. The Journal of Laboratory and Clinical Medicine, 1967, 70(3):419-428.
[29]	Schmidt P J, Toran P T, Giannetti A M, Bjorkman P J, Andrews N C. The transferrin receptor modulates Hfe-dependent regulation of hepcidin expression[J]. Cell Metabolism, 2008, 7(3):205-214.doi:10.1016/j.cmet.2007.11.016. pmid: 18316026
[30]	Yang L, Wang D, Wang X T, Lu Y P, Zhu L. The roles of hypoxia-inducible Factor-1 and iron regulatory protein 1 in iron uptake induced by acute hypoxia[J]. Biochemical and Biophysical Research Communications, 2018, 507(1/2/3/4):128-135.doi:10.1016/j.bbrc.2018.10.185. URL
[31]	Li J, Pan X H, Pan G H, Song Z J, He Y, Zhang S S, Ye X R, Yang X, Xie E J, Wang X H, Mai X D, Yin X J, Tang B Y, Shu X, Chen P Y, Dai X S, Tian Y, Yao L H, Han M L, Xu G H, Zhang H J, Sun J, Chen H, Wang F D, Min J X, Xie L W. Transferrin receptor 1 regulates thermogenic capacity and cell fate in brown/beige adipocytes[J]. Advanced Science, 2020, 7(12):1903366.doi:10.1002/advs.201903366. URL

基因 Gene	引物序列(5'-3') Primer sequences(5'-3')	产物长度/bp Product size	退火温度/℃ Annealing temperature	用途 Application
FTL	F: TTTTCCAGTCGCAACCTTCGG	664	54.9	RT-PCR
	R: GTTAGGTGGGTTCTTCAATGG
FTL	F: ACCTACCTCTCTCTGGGCTT	120	58.0	qRT-PCR
	R: CAGTTTCAAGAGACGCTCCG
GAPDH	F: AATGAAAGGGCCATCACCATC	204	59.0	qRT-PCR
	R: GTGGTTCACGCCCATCACA

基因 Gene	引物序列(5'-3') Primer sequences(5'-3')	产物长度/bp Product size	退火温度/℃ Annealing temperature	用途 Application
FTL	F: TTTTCCAGTCGCAACCTTCGG	664	54.9	RT-PCR
	R: GTTAGGTGGGTTCTTCAATGG
FTL	F: ACCTACCTCTCTCTGGGCTT	120	58.0	qRT-PCR
	R: CAGTTTCAAGAGACGCTCCG
GAPDH	F: AATGAAAGGGCCATCACCATC	204	59.0	qRT-PCR
	R: GTGGTTCACGCCCATCACA

工具Online tools	功能Analysis content	网址Website
ORF Finder	开放阅读框分析	https://www.ncbi.nlm.nih.gov/orffinder/
Blast	同源性比对分析	https://blast.ncbi.nlm.nih.gov/Blast.cgi
ProtParam	蛋白质理化性质分析	https://web.expasy.org/protparam
ProtScale	蛋白质亲疏水性分析	https://web.expasy.org/protscale
SignalP 4.1	蛋白质信号肽分析	https://services.healthtech.dtu.dk/services/SignalP-4.1/
TMHMM 2.0	蛋白质跨膜结构预测	https://services.healthtech.dtu.dk/service.php?TMHMM-2.0
NetPhos 3.1	蛋白磷酸化位点预测	https://services.healthtech.dtu.dk/service.php?NetPhos-3.1
NetNGlyc	蛋白糖基化位点预测	https://services.healthtech.dtu.dk/service.php?NetNGlyc-1.0
PSORT Ⅱ	蛋白亚细胞定位预测	https://www.genscript.com/psort.html
SOPMA	蛋白质二级结构预测	https://npsa-pbil.ibcp.fr/cgi-bin/npsa_automat.pl?page=npsa_sopma.html
STRING	蛋白互作网络预测	STRING: functional protein association networks (string-db.org)
SWISS-MODEL	蛋白质三级结构预测	https://swissmodel.expasy.org/interactive

工具Online tools	功能Analysis content	网址Website
ORF Finder	开放阅读框分析	https://www.ncbi.nlm.nih.gov/orffinder/
Blast	同源性比对分析	https://blast.ncbi.nlm.nih.gov/Blast.cgi
ProtParam	蛋白质理化性质分析	https://web.expasy.org/protparam
ProtScale	蛋白质亲疏水性分析	https://web.expasy.org/protscale
SignalP 4.1	蛋白质信号肽分析	https://services.healthtech.dtu.dk/services/SignalP-4.1/
TMHMM 2.0	蛋白质跨膜结构预测	https://services.healthtech.dtu.dk/service.php?TMHMM-2.0
NetPhos 3.1	蛋白磷酸化位点预测	https://services.healthtech.dtu.dk/service.php?NetPhos-3.1
NetNGlyc	蛋白糖基化位点预测	https://services.healthtech.dtu.dk/service.php?NetNGlyc-1.0
PSORT Ⅱ	蛋白亚细胞定位预测	https://www.genscript.com/psort.html
SOPMA	蛋白质二级结构预测	https://npsa-pbil.ibcp.fr/cgi-bin/npsa_automat.pl?page=npsa_sopma.html
STRING	蛋白互作网络预测	STRING: functional protein association networks (string-db.org)
SWISS-MODEL	蛋白质三级结构预测	https://swissmodel.expasy.org/interactive

氨基酸 Amino acid	数量/个 Amount	含量/% Content	氨基酸 Amino acid	数量/个 Amount	含量/% Content
丙氨酸Ala (A)	15	8.6	亮氨酸Leu (L)	28	16.0
精氨酸Arg (R)	13	7.4	赖氨酸Lys (K)	10	5.7
天冬酰胺Asn(N)	8	4.6	甲硫氨酸Met(M)	4	2.3
天冬氨酸Asp(D)	12	6.9	苯丙氨酸Phe(F)	8	4.6
半胱氨酸Cys(C)	1	0.6	脯氨酸Pro(P)	3	1.7
谷氨酰胺Gln(Q)	10	5.7	丝氨酸Ser(S)	7	4.0
谷氨酸Glu(E)	15	8.6	苏氨酸Thr(T)	5	2.9
甘氨酸Gly(G)	13	7.4	色氨酸Trp(W)	1	0.6
组氨酸His(H)	6	3.4	酪氨酸Tyr(Y)	5	2.9
异亮氨酸Ile(I)	3	1.7	缬氨酸Val(V)	8	4.6

Please choose a citation manager

Content to export