猪FABP1和FABP2基因的分子特征及组织表达分析

【目的】明确脂肪酸结合蛋白(FABPs)对香苏杂交猪脂肪沉积的影响,为后续进一步探究猪脂肪细胞内FABP1和FABP2基因对脂肪酸代谢的调控作用打下基础。【方法】采集香苏杂交猪和柯乐猪的心脏、肝脏、脾脏、肺脏、肾脏、大肠、小肠、背最长肌及脂肪等组织样品提取总RNA,利用PCR克隆香苏杂交猪FABP1和FABP2基因编码区(CDS)序列,通过ProtParam、ProtScale、SOPMA、SWISS-MODEL、PSORT Ⅱ Preadict及SignalP-5.0等在线软件进行生物信息学分析,并采用实时荧光定量PCR检测FABP1和FABP2基因在香苏杂交猪和柯乐猪各组织中的表达情况。【结果】香苏杂交猪FABP1和FABP2基因CDS序列分别为384和399 bp,分别编码127和132个氨基酸残基,对应的编码蛋白相对分子量为14107.23和15217.34 Da,均为亲水性蛋白;其二、三级结构均以β-折叠和无规则卷曲为主,无信号肽剪切位点,主要定位于细胞质。香苏杂交猪FABP1和FABP2氨基酸序列表现为与人类、牛、山羊的FABP1和FABP2氨基酸序列高度同源,对应的相似性均在80.0%以上,除斑马鱼外,与其他参考物种的相似性也在74.0%以上,故推测FABP1和FABP2基因序列的保守性较高;STRING预测结果显示,与这2个蛋白可能互作的蛋白有FABP5、FABP7、过氧化物酶体增殖物激活受体α(PPARα)、猪载脂蛋白A4(APOA4)、APOA1及脂酰辅酶A氧化酶1(ACOX1)等。FABP1和FABP2基因在香苏杂交猪和柯乐猪的各组织中均有表达,且以在肝脏、小肠和脂肪中的相对表达量较高,而背最长肌中的相对表达量最低;此外,FABP1和FABP2基因在香苏杂交猪肝脏和小肠中的相对表达量极显著高于其在柯乐猪对应组织中的相对表达量(P<0.01)。【结论】FABP1和FABP2基因在猪的各组织中均有表达,且以在甘油三酯和脂肪酸合成代谢的相关组织(肝脏、小肠和脂肪)中特异性高表达,可能对脂肪的合成代谢起重要调控作用。

Molecular characteristics and tissue expression analysis of FABP1 and FABP2 genes in pig

【Objective】To clarify the effect of fatty acid binding proteins(FABPs)on fat deposition in Xiangsu hybrid pigs,so as to lay the foundation for further exploring the regulatory role of FABP1 and FABP2 genes on fatty acid metabolism in adipocytes of pigs.【Method】Total RNA was extracted from the heart,liver,spleen,lung,kidney,large intestine,small intestine,longest back muscle and adipose tissue samples of Xiangsu hybrid pigs and Kole pigs,and the sequences of FABP1 and FABP2 gene coding region(CDS)of Xiangsu hybrid pigs were cloned by PCR,and were analyzed by ProtParam,ProtScale,SOPMA,SWISS-MODEL,PSORT Ⅱ Preadict and SignalP-5.0 online software for bioinformatics analysis,and RT-qPCR was used to detect the expression of FABP1 and FABP2 genes in various tissues of Xiangsu hybrid pigs and Kole pigs.【Result】CDS sequences of FABP1 and FABP2 genes of Xiangsu hybrid pigs were 384and 399 bp,encoding 127 and 132 amino acid residues,respectively,corresponding to the relative molecular weights of 14107.23 and 15217.34 Da,which are both hydrophilic proteins;their secondary and tertiary structures were dominated by β-folding and irregular coiling,with no signal peptide shear sites,and localized in the cytoplasm. The amino acid sequences of both FABP1 and FABP2 of Xiangsu hybrid pigs showed high homology with those of human,bovine and goat,and the corresponding similarities were above 80.0%,and the similarities with other reference species,except zebrafish,were also above 74.0%,so the conserved sequences of FABP1 and FABP2 genes were presumed to be high;STRING prediction results showed that the proteins that might interact with these 2 proteins were FABP5,FABP7,peroxisome proliferator-activated receptor α(PPARα),porcine apolipoprotein A4(APOA4),APOA1 and acyl-co A oxidase1(ACOX1),etc. FABP1 and FABP2 genes were expressed in all tissues of the Xiangsu hybrid pigs and Kole pigs,and were mainly expressed in various tissues,and the relative expression of FABP1 and FABP2 genes was higher in liver,small intestine and fat,while the relative expression in the longest dorsal muscle was the lowest;in addition,the relative expression of FABP1 and FABP2 genes in liver and small intestine in Xiangsu hybrid pigs was significantly higher than that in the corresponding tissues of the Kole pigs(P<0.01).【Conclusion】FABP1 and FABP2 genes are expressed in various tissues of different pigs,with specific high expression in tissues related to triglyceride and fatty acid anabolism(liver,small intestine and adipose),which may play an important role in regulating lipid anabolism.