Cloning and sequence analysis of rat fertilin alpha and beta--developmental expression, processing and immunolocalization.
Fertilin alpha and beta are members of the MDC (metalloproteinase-like, disintegrin-like, cysteine-rich) protein family and are expressed on the sperm surface where they have been proposed to play a role in mammalian fertilization. Inhibition of sperm-oocyte binding and sperm-oocyte fusion make fertilin an attractive target for the development of an immunocontraceptive vaccine. Full-length cDNAs encoding alpha and beta fertilin subunits were isolated from a rat testis cDNA library and sequenced. Using reverse transcription-polymerase chain reaction (RT-PCR), the developmental expression of fertilin alpha and beta was determined in pre-pubertal and mature rat testes. Fertilin alpha mRNA was present at all stages of development, suggesting that it is not exclusively expressed in post-meiotic germ cells. In contrast, fertilin beta mRNA was first identified in day 19 testes, coincident with the presence of pachytene spermatocytes. Polyclonal antisera raised against a 28-residue peptide (corresponding to part of the disintegrin domain) and two recombinant fusion proteins identified a 90 kDa protein in testicular sperm extracts and a 60 kDa protein in caput and cauda epididymidal sperm extracts, the predicted sizes for rat fertilin beta precursor and mature protein respectively. Indirect immunofluorescence using the anti-peptide antisera stained the acrosomal cap of permeabilized testicular, caput and caudal spermatozoa and elongating spermatids in testicular sections.
McLaughlin
E A
EA
University Department of Obstetrics and Gynaecology, St. Michael's Hospital, Bristol, UK.
Frayne
J
J
Barker
H L
HL
Jury
J A
JA
Jones
R
R
Ford
W C
WC
Hall
L
L
D051722:ADAM Proteins; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000032:analysis / Q000737:chemistry / Q000235:genetics; D000072196:Fertilins; D018507:Gene Expression Regulation, Developmental; D007150:Immunohistochemistry; D008297:Male; D008562:Membrane Glycoproteins / Q000235:genetics*; D008666:Metalloendopeptidases / Q000235:genetics*; D008969:Molecular Sequence Data; D011499:Protein Processing, Post-Translational; D051381:Rats; D017208:Rats, Wistar; D012098:Reproduction / Q000235:genetics / Q000502:physiology; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid; D013737:Testis / Q000737:chemistry; D014158:Transcription, Genetic
D018076:DNA, Complementary; D008562:Membrane Glycoproteins; D051722:ADAM Proteins; C000606456:Adam1a protein, rat; C000606459:Adam2 protein, rat; D000072196:Fertilins; D008666:Metalloendopeptidases