Localization and characterization of an orphan receptor, guanylyl cyclase-G, in mouse testis and sperm.
We recently identified a novel testis-enriched receptor guanylyl cyclase (GC) in the mouse, designated mGC-G. To further investigate its protein expression and function, we generated a neutralizing antibody specifically against the extracellular domain of this receptor. RT-PCR and immunohistochemical analyses show that mGC-G is predominantly expressed from round spermatids to spermatozoa in mouse testis at both the mRNA and protein levels. Flow cytometry and confocal immunofluorescence reveal that mGC-G is a cell surface protein restricted to the plasma membrane overlying the acrosome and midpiece of the flagellum in mature sperm. Interestingly, Western blot analysis demonstrates that testicular mGC-G is approximately 180 kDa but is subject to limited proteolysis during epididymal sperm transport, resulting in a smaller fragment tethered on the mature sperm surface. On Fluo-3 cytometrical analysis and computer-assisted sperm assay, we found that serum albumin-induced elevation of sperm intracellular Ca(2+) concentration, protein tyrosine phosphorylation, and progressive motility associated with capacitation are markedly reduced by preincubation of the anti-mGC-G neutralizing antibody. Together, these results indicate that mGC-G is proteolytically modified in mature sperm membrane and suggest that mGC-G-mediated signaling may play a critical role in gamete/reproductive biology.
Huang
Yen-Hua
YH
Department of Biochemistry and Graduate Institute of Medical Sciences, School of Medicine, Taipei Medical University, Taipei 110, Taiwan.
Wei
Chih-Chun
CC
Su
Yueh-Hsing
YH
Wu
Bo-Tsung
BT
Ciou
Yi-Yun
YY
Tu
Cheng-Fen
CF
Cooper
Trevor G
TG
Yeung
Ching-Hei
CH
Chu
Sin-Tak
ST
Tsai
Ming-Tzu
MT
Yang
Ruey-Bing
RB
D000818:Animals; D019138:Antibodies, Blocking / Q000494:pharmacology; D015153:Blotting, Western; D005434:Flow Cytometry; D006162:Guanylate Cyclase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D007136:Immunoglobulins / Q000378:metabolism; D008297:Male; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D051379:Mice; D010766:Phosphorylation; D012333:RNA, Messenger / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013081:Sperm Motility / Q000502:physiology; D013094:Spermatozoa / Q000378:metabolism*; D013737:Testis / Q000166:cytology / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism
D019138:Antibodies, Blocking; D007136:Immunoglobulins; D008565:Membrane Proteins; D012333:RNA, Messenger; D014443:Tyrosine; C487900:GC-G protein, mouse; D006162:Guanylate Cyclase