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精子蛋白质分析及精子PDC-109与公牛生育力的关联

Profiling of sperm proteins and association of sperm PDC-109 with bull fertility.

作者信息

Somashekar Lakshminarayana, Selvaraju Sellappan, Parthipan Sivashanmugam, Ravindra Janivara Parameswaraiah

机构信息

a Reproductive Physiology Laboratory, Animal Physiology Division, ICAR-National Institute of Animal Nutrition and Physiology and.

b Department of Biochemistry , Jain University , Bengaluru , India.

出版信息

Syst Biol Reprod Med. 2015;61(6):376-87. doi: 10.3109/19396368.2015.1094837. Epub 2015 Oct 20.

Abstract

The composition of sperm proteins influences the fertilizing ability of sperm and hence the present study was conducted (i) to profile sperm proteins expression patterns in bulls of differing fertility index and (ii) to identify and relate the abundant sperm proteins with bull fertility. The semen samples were collected from Holstein-Friesian bulls (n = 12) varying in conception rate (CR) (high/low). The frozen semen straws (three ejaculates, from each bull) were used to study (a) sperm kinetic parameters, (b) plasmalemma integrity, (c) mitochondrial membrane potential, and (d) chromatin distribution. Three bulls were randomly selected from each group (n = 3) and the neat sperm pellets were subjected to percoll purification, followed by protein isolation using 0.1% Triton X100. The sperm kinetic parameters, plasmalemma integrity, mitochondrial membrane potential, and the chromatin distribution did not differ significantly between groups. The number of acidic (pI; 3.1-5.6, 37%) and basic (pI; 7.9-10.0, 27%) proteins and their pattern of expression varied significantly (p < 0.05) between high and low fertile bulls. The abundant sperm protein spots in 2D-gel electrophoresis (2DE) were identified as seminal plasma protein PDC-109 (i.e., protein with N-terminus aspartic acid, D and carboxy terminus cystine, having 109 amino acids) and its isoform and spermadhesin-1 (SPADH1). The western blot analysis confirmed the presence of PDC-109 isoform proteins at 15.4 kDa (pI 5.3 and 5.5). The seminal plasma protein PDC-109 was abundant in the low fertile when compared to the high fertile group (p < 0.05). This study suggests that the imbalance in acidic and basic sperm proteins may influence sperm fertility and sperm PDC-109 levels above a certain threshold affects bull fertility.

摘要

精子蛋白质的组成会影响精子的受精能力,因此开展了本研究,(i)以剖析不同生育指数公牛的精子蛋白质表达模式,以及(ii)识别丰富的精子蛋白质并将其与公牛生育力相关联。从受孕率(CR)不同(高/低)的荷斯坦-弗里生公牛(n = 12)采集精液样本。冷冻精液细管(来自每头公牛的三次射精)用于研究(a)精子动力学参数,(b)质膜完整性,(c)线粒体膜电位,以及(d)染色质分布。从每组中随机选择三头公牛(n = 3),将纯净的精子沉淀进行Percoll纯化,随后使用0.1% Triton X100进行蛋白质分离。两组之间的精子动力学参数、质膜完整性、线粒体膜电位和染色质分布没有显著差异。高生育力和低生育力公牛之间酸性(pI;3.1 - 5.6,37%)和碱性(pI;7.9 - 10.0,27%)蛋白质的数量及其表达模式存在显著差异(p < 0.05)。二维凝胶电泳(2DE)中丰富的精子蛋白质斑点被鉴定为精浆蛋白PDC - 109(即N端为天冬氨酸、D,C端为胱氨酸,有109个氨基酸的蛋白质)及其异构体和精子黏附素 - 1(SPADH1)。蛋白质印迹分析证实了15.4 kDa(pI 5.3和5.5)处存在PDC - 109异构体蛋白。与高生育力组相比,低生育力组中精浆蛋白PDC - 109含量丰富(p < 0.05)。本研究表明,酸性和碱性精子蛋白质的失衡可能会影响精子生育力,且精子PDC - 109水平超过一定阈值会影响公牛生育力。

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