HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal My Folders Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Chayko, C. A. Articles by Tulsiani, D. R.P. Search for Related Content PubMed PubMed Citation Articles by Chayko, C. A. Articles by Tulsiani, D. R.P. Agricola Articles by Chayko, C. A. Articles by Tulsiani, D. R.P.

Biosynthesis, Processing, and Subcellular Localization of Rat Spermß-D-Galactosidase¹

^a Center for Reproductive Biology Research, ^c Department of Obstetrics & Gynecology and ^b Department of Cell Biology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-2633

ABSTRACT

During spermatogenesis, spermatids synthesize constituent proteins present in mature spermatozoa; however, little information exists on the molecular processes involved. In previous studies, this laboratory reported the characterization of rat sperm ß-D-galactosidase. In this paper, we report the localization of this enzyme along with its biosynthesis and processing. An antibody against rat luminal fluid ß-D-galactosidase was used to immunolocalize the enzyme in the testis and in epididymal spermatozoa. We found that ß-D-galactosidase is localized within the acrosomal cap of spermatids and in the acrosome and cytoplasmic droplet of epididymal spermatozoa. A combination of germ cell radiolabeling, immunoprecipitation, SDS-PAGE, and autoradiography revealed that spermatids produce two forms of ß-D-galactosidase, 90 and 88 kDa. During pulse-chase analysis, a 56-kDa form appeared. Treatment of ß-D-galactosidase immunoprecipitates from testicular spermatozoa with N-glycanase or Endo H revealed that both the 90- and 88-kDa forms become a 70-kDa polypeptide on SDS-PAGE. Since Endo H or N-glycanase treatment provided similar results, the presence of extensive N-linked high mannose/hybrid-type glycans on these proteins is indicated. Treatment of the 56-kDa form of ß-D-galactosidase with Endo H or N-glycanase resulted in the appearance of 52- and 50-kDa forms, respectively. This result suggests that the 56-kDa form contains N-linked high mannose/hybrid as well as complex oligosaccharides. During epididymal maturation, the 90-kDa form of ß-D-galactosidase persists in caput epididymal spermatozoa and is gradually converted to a major 74-kDa form in cauda spermatozoa. In addition to the 90- to 74-kDa forms, cauda spermatozoa show a 56- to 52-kDa form on Western immunoblots. Since only the high-molecular weight forms of ß-D-galactosidase are present on immunoblots of isolated sperm heads, we suggest that they are acrosomal in origin and that the 56-kDa form, which is processed to 52 kDa in cauda spermatozoa, is associated with the cytoplasmic droplet.

FOOTNOTES

First decision: 9 November 1999.

¹ Supported by National Institutes of Health grants HD25869, HD03820, and HD05797 and by an American Fellowship to C.A.C. from the American Association of University Women Educational Foundation.

² Correspondence: Marie-Claire Orgebin-Crist, Center for Reproductive Biology Research, Vanderbilt University School of Medicine, Rm. C3306 MCN, 1161 21st Ave. S., Nashville, TN 37232-2633. FAX: 615 343 7797; m-c.orgebin-crist{at}mcmail.vanderbilt.edu

This article has been cited by other articles:

				K. J. Johnson, A. Zecevic, and E. J. Kwon Protocadherin {alpha}3 Acts at Sites Distinct from Classic Cadherins in Rat Testis and Sperm Biol Reprod, February 1, 2004; 70(2): 303 - 312. [Abstract] [Full Text] [PDF]