小鼠NIS蛋白的生物信息学特征及其在乳腺中的表达分析

doi:10.16431/j.cnki.1671-7236.2020.05.003

摘要/Abstract

摘要： 为探究小鼠钠碘转运体（Na⁺/I^- symporter，NIS）蛋白的结构、功能及其在乳腺组织的表达特征，本研究扩增小鼠乳腺NIS基因编码区，测序鉴定后采用结构预测软件对NIS蛋白进行生物信息学分析，同时采用Western blotting、免疫组织化学技术和半定量PCR技术对NIS基因在不同乳腺发育时期的转录和表达水平进行分析。结果显示，小鼠NIS基因编码区全长1 857 bp，与人NIS的同源性为80.40%，保守性较强。NIS蛋白含有618个氨基酸残基，分子质量约为65.7 ku，分子式为C₂₉₉₆H₄₇₀₁N₇₄₉O₈₃₃S₃₂，理论等电点为7.44，半衰期为30 h，消光系数为102 510，不稳定系数为29.31，脂溶系数为105.91，富含疏水性氨基酸，为稳定的疏水蛋白。NIS蛋白二级结构含有α-螺旋（41.42%）、延伸链（18.28%）、β-转角（4.05%）和无规则卷曲（36.25%），三级结构与之相一致。Western blotting结果显示，4个时期均出现特异性杂交带，大小为65.7 ku，与预期结果相符。免疫组织化学结果表明，NIS蛋白在小鼠的青春期、妊娠期到泌乳期的乳腺组织中表达量逐渐增加，在泌乳期达到高峰，存在显著差异（P<0.05），在退化期逐渐恢复到类似青春期状态；NIS基因mRNA的变化规律与蛋白表达一致。本研究结果为进一步探究NIS蛋白在乳腺发育和泌乳调控中的作用提供了参考依据。

关键词: 小鼠; NIS蛋白; 乳腺; 生物信息学; 表达

Abstract: In order to explore the structure,function and expression characteristics of Na⁺/I^- symporter (NIS) protein of mammary gland in mouse,NIS gene was amplified from mammary gland in mouse,after sequencing,NIS protein was analyzed by bioinformatics software in this study.The transcription and expression level of NIS gene were detecteded in different stages of mammary gland development by Western blotting,immunohistochemistry and semi-quantitative PCR.The results showed that the CDS sequences of NIS gene was 1 857 bp,and the homology was 80.40% between Homo sapiens and Mus musculus,the conservation was stronger.There were 618 amino acid residues of NIS protein in mouse,with a molecular weight of 65.7 ku,the molecular formula was C₂₉₉₆H₄₇₀₁N₇₄₉O₈₃₃S₃₂,the theoretical isoelectric point was 7.44,the half-life was 30 h,the extinction coefficient was 102 510,the unstability coefficient was 29.31,and the fat solubility coefficient was 105.91.It was rich in hydrophobic amino acids, and was stable hydrophobin.The secondary structure of NIS protein contained alpha helix (41.42%),extended chain (18.28%),beta turn (4.05%) and random coil (36.25%),which was consist with the tertiary structural results.Western blotting results showed that the specific hybridization bands appeared in all four periods,with a size of 65.7 ku in line with expected results.The immunohistochemistry results showed that the expression of NIS protein increased gradually from puberty and pregnancy to lactation,reached the peak in lactation,there was a significant difference (P<0.05),and returned to a similar puberty state in degenerative period.The level of NIS mRNA was consistent with NIS protein.This study provided a reference for further exploring the role of NIS protein in mammary gland development and lactation regulation.

Key words: mouse; NIS protein; mammary gland; bioinformatics; expression

中图分类号:

Q786

徐笑, 景一明, 王权锜, 许立敏, 刘玉芬. 小鼠NIS蛋白的生物信息学特征及其在乳腺中的表达分析[J]. 中国畜牧兽医, 2020, 47(5): 1307-1317.

XU Xiao, JING Yiming, WANG Quanqi, XU Limin, LIU Yufen. Bioinformatics Characteristics and Expression Analysis of NIS Protein of Mammary Gland in Mouse[J]. China Animal Husbandry and Veterinary Medicine, 2020, 47(5): 1307-1317.

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