Cloning, Structure and Regulation of the sp56 Gene

Bleil博士实验室的主要兴趣是在分子水平上分析哺乳动物受精，结合细胞生物学、生物化学和分子遗传学技术。他已经鉴定出了一种小鼠精子蛋白，一种叫做sp56的凝集素，它具有许多人们所期望的蛋白质的特性，这些蛋白质负责精子识别小鼠卵细胞的透明带。他最近分离并克隆了编码sp56的cDNA。该cDNA的氨基酸序列表明：1)该蛋白在内质网上翻译，2)无跨膜结构域，3)是细胞外蛋白受体超家族的成员。针对sp56及其mRNA的探针显示，sp56基因的转录和mRNA的翻译只发生在小鼠的生精细胞中。本文所述研究的长期目标是：1)从遗传学上证明或反驳sp56是识别小鼠卵细胞透明带并介导精子与卵细胞透明带结合的精子蛋白的假设；2)确定sp56/ZP3复合物的分子结构。为了实现这些目标，必须鉴定和分离编码sp56的基因。本研究的具体目的如下：1)利用本实验室现有的编码sp56的cDNA，筛选小鼠基因组文库。基因组DNA克隆，包括翻译区上游的序列，将被纯化和测序；2)分析sp56基因序列，寻找可能编码睾丸特异性表达的调控序列。与其他先前鉴定的在睾丸中表达的基因比较，可能揭示共同的调控序列。＊＊＊

9509863 Bleil The primary interest of Dr. Bleil's laboratory is analysis of mammalian fertilization at the molecular level, combining techniques of cell biology, biochemistry and molecular genetics. He has identified a mouse sperm protein, a lectin called sp56, which has many properties expected of the protein responsible for sperm recognition of the mouse egg's zona pellucida. He has recently isolated and cloned the cDNA encoding sp56. Amino acid sequence derived from this cDNA indicates that: 1) the protein is translated on the endoplasmic reticulum, 2) has no transmembrane domain, and 3) is a member of a superfamily of extracellular protein receptors. Probes directed against sp56 and its mRNA reveal that transcription of the sp56 gene and translation of its mRNA occur exclusively in spermatogenic cells of the mouse. The long term goals to be addressed by studies described here are: 1) to genetically prove or disprove the hypothesis that sp56 is the sperm protein which recognizes the mouse egg zona pellucida, mediating sperm binding to the egg's zona pellucida, and 2) to determine the molecular structure of the sp56/ZP3 complex. In order to attain these goals, the gene encoding sp56 must be identified and isolated. The specific aims of the proposed study are as follows: 1) To use cDNA encoding sp56, currently available in this laboratory, to screen a mouse genomic library. Genomic DNA clones, including sequence upstream of the translated region, will be purified and sequenced; 2) To analyze the sequence of the sp56 gene in order to identify regulatory sequences which might encode specific expression in testes. Comparison with other, previously identified genes which are expressed in testes may reveal common regulatory sequences. ***