Johnston D S, Shaper J H, Shaper N L, Joziasse D H, Wright W W
Department of Population Dynamics, Johns Hopkins University School of Hygiene and Public Health, Baltimore, Maryland 21205-2179, USA.
Dev Biol. 1995 Sep;171(1):224-32. doi: 10.1006/dbio.1995.1273.
An essential step in murine fertilization is the binding of acrosome-intact sperm to specific O-linked glycans on zona pellucida glycoprotein 3 (ZP3). While there is agreement on the primary role of O-linked glycans in sperm-ZP3 binding, there is a striking lack of consensus on both the terminal monosaccharide(s) required for a functional binding site and the cognate protein on the sperm cell surface that recognizes this glycan. Much current debate centers on the essential role of nonreducing terminal N-acetyl-glucosaminyl or alternatively, alpha-galactosyl residues, to form a functional sperm binding ligand. Relevant to this debate, we demonstrated that alpha 1,3-galactosyltransferase (alpha 3-GT), which adds nonreducing terminal alpha-galactosyl residues to glycans, is not expressed in murine spermatocytes or spermatids. The objectives of this study were to determine whether alpha 3-GT is expressed in female germ cells and to compare the pattern of expression of two other terminal glycosyltransferases, beta 1,4-galactosyltransferase (beta 4-GT) and alpha 2,6-sialyltransferase (alpha 6-ST), between male and female germ cells. Total RNA was isolated from growing oocytes obtained from 15-day-old animals, fully grown oocytes, and eggs as well as spermatogonia, spermatocytes, and spermatids. The presence of alpha 3-GT, beta 4-GT, and alpha 6-ST mRNAs was analyzed by an RT-PCR-based assay. Our data demonstrate that the alpha 3-GT gene is expressed in female germ cells, but not in male germ cells. In contrast, both beta 4-GT and alpha 6-ST are expressed during oogenesis and spermatogenesis. This differential expression of alpha 3-GT in female germ cells is consistent with the model of sperm-egg binding in which a nonreducing terminal alpha-galactosyl residue is required for a functional determinant on ZP3 and with our hypothesis that the biological significance for the suppression of alpha 3-GT expression in male germ cells is to prevent sperm-sperm aggregation.
小鼠受精过程中的一个关键步骤是顶体完整的精子与透明带糖蛋白3(ZP3)上特定的O-连接聚糖结合。虽然对于O-连接聚糖在精子与ZP3结合中的主要作用已达成共识,但对于功能性结合位点所需的末端单糖以及精子细胞表面识别该聚糖的同源蛋白,却缺乏显著的共识。当前许多争论集中在非还原末端N-乙酰葡糖胺或α-半乳糖基残基形成功能性精子结合配体的关键作用上。与这场争论相关的是,我们证明了向聚糖添加非还原末端α-半乳糖基残基的α1,3-半乳糖基转移酶(α3-GT)在小鼠精母细胞或精子细胞中不表达。本研究的目的是确定α3-GT是否在雌性生殖细胞中表达,并比较另外两种末端糖基转移酶β1,4-半乳糖基转移酶(β4-GT)和α2,6-唾液酸转移酶(α6-ST)在雄性和雌性生殖细胞中的表达模式。从15日龄动物获取的生长中的卵母细胞、完全成熟的卵母细胞、卵子以及精原细胞、精母细胞和精子细胞中分离总RNA。通过基于逆转录聚合酶链反应(RT-PCR)的检测方法分析α3-GT、β4-GT和α6-ST mRNA的存在情况。我们的数据表明,α3-GT基因在雌性生殖细胞中表达,但在雄性生殖细胞中不表达。相反,β4-GT和α6-ST在卵子发生和精子发生过程中均有表达。α3-GT在雌性生殖细胞中的这种差异表达与精子-卵子结合模型一致,即在该模型中,ZP3上的功能性决定因素需要非还原末端α-半乳糖基残基,并且与我们的假设相符,即雄性生殖细胞中α3-GT表达受抑制的生物学意义是防止精子-精子聚集。