Delineation of regions in the extracellular domain of follicle-stimulating hormone receptor involved in hormone binding and signal transduction.
OBJECTIVE
To use antipeptide antibodies to potential surface-oriented regions of the extracellular domain (ECD) of the human follicle-stimulating hormone receptor (hFSHR) to delineate regions involved in FSH binding and FSH-induced signal transduction.


METHODS
We developed and characterized antipeptide antibodies to different, potentially surface-oriented regions of the ECD of hFSHR. The ability of these antibodies to recognize the receptor was studied by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by Western blotting. The ability to modulate FSH binding and cAMP generation was studied by the radioreceptor assay and in vitro FSH bioassay respectively.


RESULTS
Antipeptide antibodies to regions 15-31, 216-235, 285-300 and 327-341 hFSHR inhibited both FSH binding and cAMP production. Regions 15-31 and 216-235 were accessible to their cognate antipeptide antibodies both before and after FSH binding, while regions 285-300 and 327-341 hFSHR were accessible only prior to FSH binding.


CONCLUSIONS
Based on the observations made with respect to accessibility to antipeptide antibodies, ability of antibodies to inhibit FSH binding and the subsequent cAMP generation and kinetics of antibody binding, regions 285-300 and 327-341 hFSHR appear to be the chief FSH-binding sites, while regions 15-31 and 216-235 hFSHR serve as ancillary FSH-binding sites.
Kene
Pallavi S
PS
Division of Structural Biology, National Institute for Research in Reproductive Health (ICMR), Mumbai, India.
Dighe
Rajan R
RR
Mahale
Smita D
SD
D000262:Adenylyl Cyclases / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000276:immunology; D001665:Binding Sites; D002460:Cell Line; D005640:Follicle Stimulating Hormone / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D006801:Humans; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D017434:Protein Structure, Tertiary; D011817:Rabbits; D011962:Receptors, FSH / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D015398:Signal Transduction / Q000187:drug effects
D000906:Antibodies; D010446:Peptide Fragments; D011962:Receptors, FSH; D005640:Follicle Stimulating Hormone; D000262:Adenylyl Cyclases