Molecular cloning of rat sperm galactosyl receptor, a C-type lectin with in vitro egg binding activity

Abstract

Rat sperm galactosyl receptor is a member of the C‐type animal lectin family showing preferential binding to N‐acetylgalactosamine compared to galactose. Binding is mediated by a Ca2+‐dependent carbohydrate‐recognition domain (CRD) identical to that of the minor variant of rat hepatic lectin receptor 2/3 (RHL‐2/3). The molecular organization of the genomic DNA, cDNA, and derived amino acid sequence of rat testis galactosyl receptor have been determined and in vitro fertilization studies were conducted to ascertain its role. We have determined that the rat testis galactosyl receptor gene generates two mRNA species: one species, designated liver‐type, is identical to RHL‐2/3; the other, designated testis‐type, contains one unspliced intron (86 nt) which alters the reading frame and changes the amino acid sequence of the carboxyl terminus. As a result, the CRD (glutamine‐proline‐aspartic acid/QPD) and flanked Ca2+‐binding amino acid sequences were not present in the testis‐type protein. Northern and Southern blots demonstrated presence of transcripts with unspliced intron in rat sperm but not liver. Similarly, antibody, raised against a synthetic 12‐amino acid peptide (p12) encoded by the unspliced intron, recognized in immunoblots a 54 kDa receptor protein in protein extracts from testis but not from liver. Immunofluorescence and immunogold electron microscopy studies demonstrated that both protein species localized on the plasma membrane surface of the head and tail of rat sperm. Furthermore, capacitated rat sperm preincubated with polyclonal antisera to RHL‐2/3 or to the CRD of the liver‐type galactosyl receptor showed a statistically significant decrease in the in vitro fertilization rate. We conclude that rat sperm galactosyl receptor may play a role in egg binding and that an undetermined molecular mechanism operates to generate two proteins with identical intracellular amino terminal domain but only one of them displays a CRD and associated Ca2+‐binding sites at the carboxyl terminal extracellular domain. Mol. Reprod. Dev. 56:401–411, 2000. © 2000 Wiley‐Liss, Inc.

Purification of N-acetyllactosamine-binding Activity from the Porcine Sperm Membrane: Possible Involvement of an ADAM Complex in the Carbohydrate-binding Activity of Sperm

“…Murine sperm galactosyltransferase has been the most extensively studied carbohydrate-binding protein [14], but it appears unlikely that it participates in the initial gamete adhesion process [15]. Other carbohydrate-binding proteins have been implicated, i.e., rat galactosyl receptor, which is identical to the rat hepatic lectin receptor 2/3 [16], porcine AWN-1, which interacts with core 1 O-glycans [17], and human fucosyltransferase, which binds to glycodelin-A as well as to the ZP [18]; none of these, however, is widely accepted as a sperm carbohydrate-binding protein involved in ZP binding. We previously characterized porcine ZP oligosaccharides [2,3,5].…”

Zona-Binding Inhibitory Factor-1 from Human Follicular Fluid Is an Isoform of Glycodelin1

“…A Ca 2ϩ -dependent lectin galactosyl receptor is present on the sperm membrane in some animals [41,42] and in humans [43]. This receptor binds preferentially to N-acetylgalactosamine rather than to galactose and has been suggested to play a role in spermatozoa-ZP binding [44]. This suggestion is consistent with the observation that N-acetylgalactosamine treatment inhibits the binding of hamster spermatozoa to ZP [45].…”

The C‐type lectin‐like domain superfamily

“…One of first C-type lectins discovered [10,125]. Rat spermatogenic cells express unusual ASGR2 oligomer (sperm galactosyl receptor), consisting of a full-length and a truncated form lacking C-terminal part of CTLD [18,126].…”