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Abstract

Both SMAD2 and SMAD3 mediate activin-stimulated expression of the follicle-stimulating hormone β subunit in mouse gonadotrope cells 
Bernard,Daniel J.
Molecular Endocrinology 18(3): 606-623
Publication date: 2004


The activins are pleiotropic members of the TGFß superfamily.Within the anterior pituitary gland, activins stimulate FSHsynthesis in an autocrine/paracrine fashion by stimulating transcriptionof the FSHß subunit gene. Here, the mechanisms mediatingthis effect were investigated in the murine gonadotrope cellline, LßT2. Recombinant activin A and activin B dose-and time-dependently stimulated endogenous FSHß mRNAexpression. FSHß primary transcript and mRNA levelswere increased within 30-60 min, but these effects wereblocked by preincubation with the transcription inhibitor actinomycin-D,suggesting that the FSHß gene is a direct target ofthe activin signal transduction cascade. In other systems, activinsignals are transduced through a heteromeric serine/threoninereceptor complex, which includes the signaling activin typeIB receptor [activin receptor-like kinase 4 (ALK4)]. Transfectionof a constitutively active form of the receptor, ALK4T206D,stimulated FSHß mRNA levels. Overexpression of theinhibitory SMAD7 blocked this effect, as well as activin-stimulatedFSHß transcription. Because SMAD7 functions by preventingaccess of SMAD2 and SMAD3 to ALK4, these data suggested thatboth activins and ALK4 require SMAD2 and/or SMAD3 to affectFSHß transcription. Consistent with this idea, activinA stimulated SMAD2 and SMAD3 phosphorylation and nuclear translocationwithin 5-10 min in LßT2 cells. Transient transfectionof SMAD3, but not SMADs 1, 2, 4, 5, or 8, stimulated endogenousFSHß mRNA levels. The results of SMAD2 transfectionstudies were inconclusive, however, because of a persistentfailure to overexpress the full-length SMAD2 protein specificallyin LßT2 cells. To assess more directly roles for bothSMAD2 and SMAD3 in activin-stimulated FSHß expression,RNA interference was used to decrease endogenous SMAD proteinlevels in LßT2 cells. Activin A- and ALK4T206D-stimulatedtranscription of the FSHß gene were significantlyattenuated by the depletion of either SMAD2 or SMAD3. Collectively,these data suggest that activins use both SMAD2- and SMAD3-dependentmechanisms to stimulate FSHß transcription in mousegonadotrope cells.