努比亚山羊UCP1基因克隆、生物信息学分析及组织表达研究

doi:10.16431/j.cnki.1671-7236.2023.02.002

Abstract

Abstract: 【Objective】 This study was aimed to clone uncoupling protein-1 (UCP1) gene in Nubian goats and carried out bioinformatics analysis,and detect its expression difference of UCP1 gene in different tissues of Nubian goats,in order to provide data for studying its regulatory function in fat metabolism and analyzing the function of UCP1 gene in Nubian goats.【Method】 Using the cDNA from the subcutaneous fat tissue in Nubian goats as a template,the CDS region of UCP1 gene in Nubian goats was amplified by PCR and cloned,sequence similarity alignment and phylogentic tree contruction were carried out with other species,and the bioinformatics analysis of UCP1 protein was carried out.The expression of UCP1 gene in heart,liver,spleen,kidney,longissimus dorsi muscle,subcutaneous fat and abdominal fat of Nubian goats were detected by Real-time quantitative PCR.【Result】 The CDS sequence of UCP1 gene in Nubian goats was 918 bp and encoded 305 amino acids.The similarity of amino acid sequence of UCP1 gene between Nubian goats and Ovis aries, Bos indicus× Bos taurus, Bubalus bubalis, Oryx dammah, Cervus elaphus, Canelus ferus, Equus asinus, Aliuropoda melanoleuca and Homo sapiens were 98.1%,97.0%,96.5%,96.1%,95.8%,91.0%,87.0%,86.5% and 83.8%,respectively.The phylogenetic tree indicated that the closest was to Ovis aries and the furthest was to Homo sapiens.The results of bioinformatics analysis showed that the molecular mass of UCP1 protein in Nubian goats was 32.97 ku,with an isoelectric point of 9.29,which was a basic proteins.UCP1 protein lacked stability,an overall average hydrophilicity was 0.20,which possessed certain hydrophilicity.The lipid solubility coefficient was 91.70,the predicted value of the number of amino acid residues in transmembrane helices was 15.94,the predicted value of the number of transmembrane helices in the first 60 amino acids of the protein was 8.76,the overall probability of being located on the cytoplasmic side of the membrane was 29.03%,and it belonged to nonsecretory proteins.The secondary structure was composed of alpha helix (48.85%),random coil (27.54%),extended chain (16.39%),and beta turn (7.21%).UCP1 had a total of 28 phosphate sites,5 potential O-glycosylation sites and 2 N-glycosylation potential sites.Real-time quantitative PCR results showed that the expression of UCP1 gene in subcutaneous fat of Nubian goats was relatively high,which was significantly higher than other tissues (P＜0.05),while the expression of longissimus dorsi muscle in Nubian goats was the lowest.【Conclusion】The CDS sequence of UCP1 gene was successfully amplified,UCP1 was a basic protein lacking stability and hydrophilicity. UCP1 gene was widely expressed in various tissues of Nubian goats, which was mainly expressed in subcutaneous fat and spleen.The results provided a theoretical basis for subsequent research on the mechanism of UCP1 gene in the energy supply of fat thermogenesis.

Key words: Nubian goats; UCP1 gene; cloning; bioinformatics analysis; tissue expression

CLC Number:

WEI Yirong, ZHENG Zihua, ZHANG Sanbao, SONG Ying, JIANG Haiyu, SUN Wenyue, CHENG Pengfei, LIU Yufan, ZOU Jianwei, HUANG Yanna, PAN Yan, JIANG Qinyang. Cloning,Bioinformatics Analysis and Tissue Expression of UCP1 Gene in Nubian Goats[J]. China Animal Husbandry and Veterinary Medicine, 2023, 50(2): 440-450.

References

[1] LUEVANO-MARTINEZ L A.Uncoupling proteins (UCP) in unicellular eukaryotes:True UCPs or UCP1-like acting proteins?[J]. FEBS Letters, 2012,586(7):1073-1078.
[2] ADAMS S H.Uncoupling protein homologs:Emerging views of physiological function[J]. Journal of Nutrition, 2000,132(12):711-714.
[3] NEDERGAARD J,CANNON B.Brown adipose tissue:Development and function[J]. Fetal and Neonatal Physiology (Third Edition),2004,5(10):404-415.
[4] AKYOL M,RAMAGE L,FLETCHER A,et al.Glucocorticoids acutely increase brown adipose tissue activity in humans,revealing species-specific differences in UCP-1 regulation[J]. Cell Metabolism, 2016,24(1):130-141.
[5] GURA T.Cell biology-uncoupling proteins provide new clue to obesity’s causes[J]. Science, 1998,280(5368):1369-1970.
[6] MENG Q,CHENG Y,WANG C,et al.Leucine deprivation decreases fat mass by stimulation of lipolysis in white adipose tissue and upregulation of uncoupling protein 1 (UCP1) in brown adipose tissue[J]. Diabetes, 2010,59(1):17-25.
[7] KAZAK L,CHOUCHANI E T,STAVROVSKAYA I G,et al.UCP1 deficiency causes brown fat respiratory chain depletion and sensitizes mitochondria to calcium overload-induced dysfunction[J]. Proceedings of the National Academy of Sciences of the United States of America, 2017,114(30):7981-7986.
[8] 孙国权,高树新,吴慧光,等.解偶联蛋白1、2和3基因在中国西门塔尔牛组织器官中的表达水平及其与胴体品质关系分析[J].华北农学报,2014,29(4):116-120. SUN G Q,GAO S X,WU H G,et al.Analyze the UCP1, UCP2, UCP3 genes expression level in tissues/organs and the relationship with the carcass traits in Chinese Simmental cattle[J]. Acta Agriculturae Boreali-Sinica, 2014,29(4):116-120.(in Chinese)
[9] 孙斌,唐琳,张军芳,等.延边牛 UCP1基因CDS区克隆,生物信息学分析及组织mRNA表达[J].华北农学报,2021,36(6):211-218. SUN B,TANG L,ZHANG J F,et al.Clone,bioinformatics analysis and tissue mRNA expression of Yanbian cattle UCP1 gene CDS region[J]. Acta Agriculturae Boreali-Sinica,2021,36(6):211-218.(in Chinese)
[10] LEE Y,WILLERS C,KUNJI E R S,et al.Uncoupling protein 1 binds one nucleotide per monomer and is stabilized by tightly bound cardiolipin[J]. Proceedings of the National Academy of Sciences of the United States of America, 2015,112(22):6973-6978.
[11] ABD EIDAIM M A,HASHIMOTO O,OHTSUKI H,et al.Expression of uncoupling protein 1 in bovine muscle cells [J]. Journal of Animal Science, 2016,94(12):5097.
[12] 王翠丽,林亚秋,岳宇,等.牦牛解偶联蛋白-3基因的克隆测序和肌肉表达谱[J].西北农业学报,2012,21(5):18-21. WANG C L,LIN Y Q,YUE Y,et al.Cloning and sequencing and muscle expression profile of uncoupling protein 3 in yak[J]. Acta Agriculturae Boreali-Occidentalis Sinica,2012,21(5):18-21.(in Chinese)
[13] YUAN Y N,LIU W Z,LIU J H,et al.Cloning and ontogenetic expression of the uncoupling protein 1 gene UCP1 in sheep[J]. Journal of Applied Genetics, 2012,53(2):203-212.
[14] 王秋惠,徐星科,汪夏,等.黄连碱调节 UCP1基因表达改善肥胖的分子机制研究[J].中国药学杂志,2021,56(21):1712-1721. WANG Q H,XU X K,WANG X,et al.Coptisine regulates the expression of UCP1 gene to ameliorate obesity and its associated molecular mechanisms[J]. Journal of the Chinese Pharmaceutical Association, 2021,56(21):1712-1721.(in Chinese)
[15] OKA T,LKHAGVASUREN B,NAKAMURA Y,et al.Social defeat stress induces hyperthermia through activation of thermoregulatory sympathetic premotor neurons in the medullary raphe region[J]. Neuroscience Research, 2011,71:161.
[16] KATAOKA N,HIOKI H,KANEKO T,et al.Psychological stress activates a dorsomedial hypothalamus-medullary raphe circuit driving brown adipose tissue thermogenesis and hyperthermia[J]. Cell Metabolism, 2014,20(2):346-358.
[17] JACOBSSON A,ENERBCK S,SIMPSON E M,et al.Mice lacking mitochondrial uncoupling protein are cold-sensitive but not obese see comments[J]. Nature, 1997,387(6628):90-94.
[18] TSUBOTA A,OKAMATSU-OGURA Y,BARIUAN J V,et al.Role of brown adipose tissue in body temperature control during the early postnatal period in Syrian hamsters and mice[J]. Journal of Veterinary Medical Science, 2019,81(10):1461-1467.
[19] INOKUMA K,OKAMATSU-OGURA Y,OMACHI A,et al.Indispensable role of mitochondrial UCP1 for antiobesity effect of beta 3-adrenergic stimulation[J]. American Journal of Physiology-Endocrinology and Metabolism, 2006,290(5):E1014-E1021.
[20] SAITO M,MATSUSHITA M,YONESHIRO T,et al.Brown adipose tissue,diet-induced thermogenesis,and thermogenic food ingredients:From mice to men[J]. Frontiers in Endocrinology, 2020,11:222.
[21] SAITO M,OKAMATSU-OGURA Y,MATSUSHITA M,et al.High incidence of metabolically active brown adipose tissue in healthy adult humans:Effects of cold exposure and adiposity[J]. Diabetes, 2009,58(7):1526-1531.
[22] RODRIGUEZ-CUENCA S,COCHEMÉ H M,LOGAN A,et al.Consequences of long-term oral administration of the mitochondria-targeted antioxidant MitoQ to wild-type mice[J]. Free Radical Biology and Medicine, 2010,48(1):161-172.
[23] FINKEL T.From sulfenylation to sulfhydration:What a thiolate needs to tolerate[J]. Science Signaling, 2012,5(215):10.
[24] RILEY C L,BOUTZ D,BEAN C,et al.UCP1 is essential for mitochondrial structural integrity and function in brown adipose tissue[J]. FASEB Journal, 2017,31:1230-1236.
[25] STIER A,BIZE P,HABOLD C,et al.Mitochondrial uncoupling prevents cold-induced oxidative stress:A case study using UCP1 knockout mice[J]. Journal of Experimental Biology, 2014,217(4):624-630.
[26] ZHENG Q T,LIN J,HUANG J J,et al.Reconstitution of UCP1 using CRISPR/Cas9 in the white adipose tissue of pigs decreases fat deposition and improves thermogenic capacity[J]. Proceedings of the National Academy of Sciences of the United States of America, 2017,114(45):9474-9482.
[27] PECQUEUR C,ALVES-GUERRA M C,GELLY C,et al.Uncoupling protein 2 in vivo distribution,induction upon oxidative stress,and evidence for translational regulation[J]. Journal of Biological Chemistry, 2001,276(12):8705-8712.
[28] ARSENIJEVIC D,ONUMA H,PECQUEUR C,et al.Disruption of the uncoupling protein-2 gene in mice reveals a role in immunity and reactive oxygen species production[J]. Nature Genetics, 2000,26(4):435-439.
[29] FLACHS P,SPONAROVA J,KOPECKY P,et al.Mitochondrial uncoupling protein 2 gene transcript levels are elevated in maturating erythroid cells[J]. FEBS Letters, 2007,581(6):1093-1097.
[30] 刘建华,乔利英,郭云雁,等.绵羊 UCP2基因的克隆及其在脂肪组织中的季节性差异表达[J].畜牧兽医学报,2015,46(7):1114-1121. LIU J H,QIAO L Y,GUO Y Y,et al.Cloning of ovine UCP2 gene and its seasonally differential expression in adipose tissues[J]. Chinese Journal of Animal and Veterinary Sciences, 2015,46(7):1114-1121.(in Chinese)
[31] 赵丹,熊燕,华永琳,等.牦牛 UCP1基因生物学特性及组织表达分析[J].西南民族大学学报(自然科学版),2021,47(2):139-148. ZHAO D,XIONG Y,HUA Y L,et al.Biological characteristic and tissue expression analysis of UCP1 gene in yak(Bos grunniens)[J]. Journal of Southwest Minzu University(Natural Science Edition), 2021,47(2):139-148.(in Chinese)
[32] MARK P,HENRY B A,MARK B,et al.Ontogeny and thermogenic role for sternal fat in female sheep[J]. Endocrinology, 2017,158(7):2212-2225.
[33] SIDOSSIS L,KAJIMURA S.Brown and beige fat in humans:Thermogenic adipocytes that control energy and glucose homeostasis[J]. Journal of Clinical Investigation, 2015,125(2):478-486.
[34] CINTI S.Reversible physiological transdifferentiation in the adipose organ[J]. Proceedings of the Nutrition Society, 2009,68(4):340-349.

Cloning,Bioinformatics Analysis and Tissue Expression of UCP1 Gene in Nubian Goats

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