Identification of the gene for the developmentally expressed 70 kDa heat-shock protein (P70) of mouse spermatogenic cells.
Mouse spermatogenic cells synthesize a 70-kDa protein (P70) closely related to the major heat-shock protein (hsp70) of mammalian cells (R. L. Allen, D. A. O'Brien, and E. M. Eddy, Mol. Cell. Biol. 8, 828-832, 1988). Expression of P70 is developmentally regulated while hsp70 is induced in response to stress, suggesting that P70 is the product of a unique member of the Hsp70 multigene family transcribed in spermatogenic cells. A strong candidate for this gene was the Hsp70.2 gene (Z. F. Zakeri, D. J. Wolgemuth, and C. R. Hunt, Mol. Cell. Biol. 8, 2925-2932, 1988). A DNA segment from the 5' region of Hsp70.2 hybridized to a 2.7-kb transcript with a temporal pattern of expression in mouse spermatogenic cells similar to the P70 protein. We used a polyclonal antiserum generated against a synthetic peptide predicted from the Hsp70.2 sequence to characterize its protein product and to isolate cDNA clones from a pachytene spermatocyte expression library. The antiserum reacted specifically with meiotic and postmeiotic spermatogenic cells on sections of mouse testis. It recognized the P70 protein on Western blots of two-dimensional gels and did not bind to other heat-shock proteins of spermatogenic or somatic cells. The cDNAs hybridized to a 2.7-kb mRNA that was abundant in unstressed pachytene spermatocytes and round spermatids but was not detected in other cell types. Two cDNAs were sequenced and found to be 99% homologous to the 3' end of the Hsp70.2 gene. These data strongly supported the hypothesis that P70 is the expressed product of the Hsp70.2 gene in mouse spermatogenic cells.
Rosario
M O
MO
Gamete Biology Section, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709.
Perkins
S L
SL
O'Brien
D A
DA
Allen
R L
RL
Eddy
E M
EM
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D001483:Base Sequence; D002454:Cell Differentiation / Q000235:genetics; D015342:DNA Probes; D015723:Gene Library; D006360:Heat-Shock Proteins / Q000032:analysis / Q000235:genetics*; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D010452:Peptide Biosynthesis; D010449:Peptide Mapping; D010455:Peptides / Q000276:immunology; D012670:Seminiferous Epithelium / Q000166:cytology / Q000378:metabolism*; D016415:Sequence Alignment; D013094:Spermatozoa / Q000378:metabolism
D000906:Antibodies; D015342:DNA Probes; D006360:Heat-Shock Proteins; D010455:Peptides