Deletion of follicle-stimulating hormone (FSH) receptor residues encoded by exon one decreases FSH binding and signaling in the rat.
The rat FSH receptor (rFSHR) shares considerable homology with the rat LH receptor (rLHR), yet binds human FSH (hFSH) with high fidelity, suggesting that the binding determinant encoded by the rFSHR gene shares no homology with the analogous rLHR primary sequence, thereby affording specificity of ligand binding. Two such regions of primary sequence have been previously identified and studied by peptide challenge tests and immunoneutralization studies. We therefore implemented site-directed mutagenesis to delete the regions S9-N30 and D300-F315 of the mature rFSHR sequence. The mutant receptor (DeltarFSHR) cDNAs were expressed in insect cells. The large deletion DeltarFSHRS9-N30 and a smaller deletion, DeltarFSHRS9-S18, did not bind (125)I-hFSH. However, DeltarFSHRK19-R29 and DeltarFSHRD300-F315 bound (125)I-hFSH with an affinity indistinguishable from wild-type rFSHR. The deletion mutants DeltarFSHR S9-N30 or DeltarFSHRS9-S18 were not detectable on the cell surface by flow cytometry unless cells were sheared. Although (125)I-hFSH binding to DeltarFSHRK19-R29 was normal, this form of the receptor was defective for signal transduction whereas DeltarFSHRD300-F315 was not. Furthermore, neither region seems to be a specificity determinant, since their removal did not result in high-affinity binding of hCG to DeltarFSHR.
Mann
K I
KI
Department of Biological Sciences, State University of New York at Albany, Albany, New York 12208, USA.
Liu
X
X
Dias
J A
JA
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D015153:Blotting, Western; D002462:Cell Membrane; D002478:Cells, Cultured; D006063:Chorionic Gonadotropin / Q000378:metabolism; D005091:Exons; D005434:Flow Cytometry; D005640:Follicle Stimulating Hormone / Q000378:metabolism*; D006801:Humans; D007313:Insecta / Q000166:cytology; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D051381:Rats; D011962:Receptors, FSH / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion; D015398:Signal Transduction
D006063:Chorionic Gonadotropin; D011962:Receptors, FSH; D011994:Recombinant Proteins; D005640:Follicle Stimulating Hormone