Ma Guojian, Huang Jing, Sun Nunu, Liu Xiangdong, Zhu Mengjin, Wu Zhenfang, Zhao Shuhong
Key Lab of Agricultural Animal Genetics, Breeding, and Reproduction of Ministry of Education & Key Laboratory of Swine Genetics and Breeding of Ministry of Agriculture, Huazhong Agricultural University, Wuhan, Hubei, PR China.
Mol Immunol. 2008 May;45(10):2797-807. doi: 10.1016/j.molimm.2008.02.007. Epub 2008 Mar 17.
Experimental evidence indicated that interferon-inducible guanylate-binding proteins (GBPs) are similar with genes in the myxovirus (Mx) resistance protein subfamily of the large GTPases protein family and play important roles in the resistance to intracellular pathogens. As more diseases exert significant influence on pig industry, it is anticipated that more candidate disease-resistance genes could be found in future strategies aimed at improving genetic resistance to infectious diseases. In this study, we cloned cDNA sequences and analyzed the genomic structure of porcine GBP1 (poGBP1) and GBP2 (poGBP2). The two genes were mapped to SSC4q21-q23 and SSC4q24 by the SCHP panel respectively, further IMpRH panel analysis showed both genes were most closely linked to the marker SWR153. The deduced amino acid sequences of these two genes share the same three classical GTP-binding motifs at the amino terminus and are less conserved at the carboxyl termini except for a CaaX motif, compared with human and mouse counterparts. The reverse transcriptase-polymerase chain reaction (RT-PCR) revealed that poGBP1 and poGBP2 were both widely expressed in many tissues, and transient transfection indicated that poGBP1 and poGBP2 proteins were both located in cytoplasms within Pig Kidney Epithelial cells (PK15). Quantitative real-time PCR (Q-RT-PCR) analyses showed poGBP1 and poGBP2 had very similar expression patterns in PK15 cells at different time points after poly I:C stimulation, suggesting that ISRE (interferon-stimulated response element) plays a crucial role in the transcriptional regulation of these two genes. Four single nucleotide polymorphisms (SNPs) and three SNPs were detected in the cDNA sequences of poGBP1 and poGBP2, respectively. Association analyses revealed that the poGBP1 Eco81I and poGBP2 SspI polymorphisms both had significant associations (p<0.05) with red blood cell count (RBC), haemoglobin concentration (HGB) and hematocrit (HCT) of 17-day-old pigs.
实验证据表明，干扰素诱导的鸟苷酸结合蛋白（GBPs）与大GTP酶蛋白家族的黏液病毒（Mx）抗性蛋白亚家族中的基因相似，并在抵抗细胞内病原体方面发挥重要作用。随着越来越多的疾病对养猪业产生重大影响，预计在未来旨在提高对传染病遗传抗性的策略中会发现更多候选抗病基因。在本研究中，我们克隆了猪GBP1（poGBP1）和GBP2（poGBP2）的cDNA序列并分析了其基因组结构。这两个基因分别通过SCHP面板定位到SSC4q21-q23和SSC4q24，进一步的IMpRH面板分析表明这两个基因都与标记SWR153紧密连锁。与人类和小鼠的对应物相比，这两个基因推导的氨基酸序列在氨基末端具有相同的三个经典GTP结合基序，除了一个CaaX基序外，在羧基末端的保守性较低。逆转录聚合酶链反应（RT-PCR）显示poGBP1和poGBP2在许多组织中均广泛表达，瞬时转染表明poGBP1和poGBPigB2蛋白均位于猪肾上皮细胞（PK15）的细胞质中。定量实时PCR（Q-RT-PCR）分析表明，在聚肌胞苷酸（poly I:C）刺激后的不同时间点，poGBP1和poGBP2在PK15细胞中的表达模式非常相似，这表明干扰素刺激反应元件（ISRE）在这两个基因的转录调控中起关键作用。在poGBP1和poGBP2的cDNA序列中分别检测到四个单核苷酸多态性（SNP）和三个SNP。关联分析显示，poGBP1的Eco81I和poGBP2的SspI多态性均与17日龄仔猪的红细胞计数（RBC）、血红蛋白浓度（HGB）和血细胞比容（HCT）有显著关联（p<0.05）。
