Inhibin and inhibin alpha-chain precursors are produced by immature rat Sertoli cells in culture.
Stimulation of Sertoli cells from immature rats with dibutyryl cyclic (dbc) AMP resulted in a decrease in the ratio of inhibin biological (B):immunological (I) activities in vitro. To establish the basis for this decrease, culture medium from Sertoli cells stimulated with dbcAMP was fractionated by dye-affinity chromatography, reverse-phase HPLC, and preparative PAGE. Two peaks of inhibin activity were identified: a predominantly bioactive 29-kDa peak I material (B:I ratio = 5.0) and a bio-inactive, immunoactive 27-kDa peak II material (B:I ratio = 0.1). Evidence of a subunit structure was established by iodination and immunopurification using an inhibin alpha-subunit antiserum. On reduction, peak I (29-kDa) material showed bands of 19 kDa and 14 kDa, whereas peak II (27-kDa) material showed a single 20-kDa band. On the basis of HPLC retention position, molecular mass, evidence of subunit structures and their molecular masses, and inhibin in vitro bio- and immunoactivities, peak I and II materials were most likely inhibin and the alpha-subunit precursor protein pro-alpha C. Western blotting of Sertoli cell culture medium using antiserum directed against the NH2 terminal region (alpha N) of the alpha-subunit precursor also indicated the presence of 24-kDa alpha N. It is concluded that after dbcAMP stimulation, Sertoli cells produce 29-kDa inhibin and the alpha-subunit precursor proteins pro-alpha C and alpha N. The production of the alpha-subunit precursor in addition to inhibin provides an explanation for the decrease in the inhibin B:I ratio following dbcAMP stimulation of Sertoli cells in culture.
Hancock
A D
AD
Department of Anatomy, Monash University, Clayton, Victoria, Australia.
Robertson
D M
DM
de Kretser
D M
DM
D000818:Animals; D003994:Bucladesine / Q000494:pharmacology; D002454:Cell Differentiation; D002478:Cells, Cultured; D007265:Inhibins / Q000096:biosynthesis* / Q000302:isolation & purification; D008297:Male; D008970:Molecular Weight; D011498:Protein Precursors / Q000096:biosynthesis* / Q000302:isolation & purification; D051381:Rats; D012708:Sertoli Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*
D011498:Protein Precursors; D007265:Inhibins; D003994:Bucladesine