Novel alternatively spliced exon in the extracellular ligand-binding domain of the pituitary adenylate cyclase-activating polypeptide (PACAP) type 1 receptor (PAC1R) selectively increases ligand affinity and alters signal transduction coupling during spermatogenesis.
The expression of the paracrine signaling hormone pituitary adenylate cyclase-activating polypeptide (PACAP) is regulated in a cyclical fashion during the 12-day spermatogenic cycle of the adult rat testis. The precise functions of PACAP in the development of germ cells are uncertain, but cycle- and stage-specific expression may augment cAMP-regulated gene expression in germ cells and associated Sertoli cells. Here we report the existence of a heretofore unrecognized exon in the extracellular domain of the PACAP type 1 receptor (PAC1R) that is alternatively spliced during the spermatogenic cycle in the rat testis. This splice variant encodes a full-length receptor with the insertion of an additional 72 base pairs encoding 24 amino acids (exon 3a) between coding exons 3 and 4. The PAC1R(3a) mRNA is preferentially detected in seminiferous tubules and is expressed at the highest levels in round spermatids and Sertoli cells. Analyses of ligand binding and signaling functions in stably transfected HEK293 cells expressing the two receptor isoforms reveals a 6-fold increase in the affinity of the PAC1R(3a) to bind PACAP-38, and alterations in its coupling to both cAMP and inositol phosphate signaling pathways relative to the wild type PAC1R. These findings suggest that the extracellular region between coding exons 3 and 6 of PAC1R may play an important role in the regulation of the relative ligand affinities and the relative coupling to G(s) (cAMP) and G(q) (inositol phosphates) signal transduction pathways during spermatogenesis.
Laboratory of Molecular Endocrinology, Massachusetts General Hospital, Howard Hughes Medical Institute, Harvard Medical School, Boston, Massachusetts 02114, USA.
P B
PB
Daniel
T J
TJ
Kieffer
C A
CA
Leech
J F
JF
Habener
D017398:Alternative Splicing; D000818:Animals; D001665:Binding Sites; D001921:Brain / Q000502:physiology; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D000242:Cyclic AMP / Q000378:metabolism; D005091:Exons; D019204:GTP-Binding Proteins / Q000378:metabolism; D014644:Genetic Variation; D006801:Humans; D007295:Inositol Phosphates / Q000378:metabolism; D007515:Islets of Langerhans / Q000502:physiology; D008024:Ligands; D008297:Male; D009479:Neuropeptides / Q000737:chemistry / Q000378:metabolism; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; D010902:Pituitary Gland / Q000502:physiology; D016133:Polymerase Chain Reaction; D021122:Protein Subunits; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D051236:Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide; D051237:Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide, Type I; D011979:Receptors, Pituitary Hormone / Q000737:chemistry / Q000235:genetics* / Q000502:physiology*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012671:Seminiferous Tubules / Q000502:physiology; D015398:Signal Transduction / Q000502:physiology; D013091:Spermatogenesis / Q000502:physiology*; D013737:Testis / Q000166:cytology / Q000502:physiology*; D014158:Transcription, Genetic; D014162:Transfection
C495401:ADCYAP1 protein, human; C495403:Adcyap1 protein, rat; D007295:Inositol Phosphates; D008024:Ligands; D009479:Neuropeptides; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; D021122:Protein Subunits; D012333:RNA, Messenger; D051236:Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide; D051237:Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide, Type I; D011979:Receptors, Pituitary Hormone; D011994:Recombinant Proteins; D000242:Cyclic AMP; D019204:GTP-Binding Proteins; D002118:Calcium