Both SMAD2 and SMAD3 mediate activin-stimulated expression of the follicle-stimulating hormone β subunit in mouse gonadotrope cells

Document Type

Article (peer-reviewed)

Publication Date



The activins are pleiotropic members of the TGFβ superfamily. Within the anterior pituitary gland, activins stimulate FSH synthesis in an autocrine/ paracrine fashion by stimulating transcription of the FSHβ subunit gene. Here, the mechanisms mediating this effect were investigated in the murine gonadotrope cell line, LβT2. Recombinant activin A and activin B dose- and time-dependently stimulated endogenous FSHβ mRNA expression. FSHβ primary transcript and mRNA levels were increased within 30-60 min, but these effects were blocked by preincubation with the transcription in-hibitor actinomycin-D, suggesting that the FSHβ gene is a direct target of the activin signal transduction cascade. In other systems, activin signals are transduced through a heteromeric serine/threonine receptor complex, which includes the signaling activin type IB receptor [activin receptor-like kinase 4 (ALK4)]. Transfection of a constitutively active form of the receptor, ALK4T206D, stimulated FSHβ mRNA levels. Overexpression of the inhibitory SMAD7 blocked this effect, as well as activin-stimulated FSHβ transcription. Because SMAD7 functions by preventing access of SMAD2 and SMAD3 to ALK4, these data suggested that both activins and ALK4 require SMAD2 and/or SMAD3 to affect FSHβ transcription. Consistent with this idea, activin A stimulated SMAD2 and SMAD3 phosphorylation and nuclear translocation within 5-10 min in LβT2 cells. Transient fransfection of SMAD3, but not SMADs 1, 2, 4, 5, or 8, stimulated endogenous FSHβ mRNA levels. The results of SMAD2 transfection studies were inconclusive, however, because of a persistent failure to overexpress the full-length SMAD2 protein specifically in LβT2 cells. To assess more directly roles for both SMAD2 and SMAD3 in activin-stimulated FSHβ expression, RNA interference was used to decrease endogenous SMAD protein levels in LβT2 cells. Activin A- and ALK4T206D-stimulated transcription of the FSHβ gene were significantly attenuated by the depletion of either SMAD2 or SMAD3. Collectively, these data suggest that activins use both SMAD2- and SMAD3-dependent mechanisms to stimulate FSHβ transcription in mouse gonadotrope cells.