The 3 Untranslated Region of Bovine Follicle-Stimulating Hormone (beta) Messenger RNA Downregulates Reporter Expression: Involvement of AU-Rich Elements and Transfactors

FSH(beta) mRNA has a unique 3ʹ untranslated region (3ʹUTR) that is highly conserved across the species. Sequence analyses of the mouse, rat, human, bovine, and ovine 3ʹUTRs revealed the presence of elements implicated in mRNA instability and translational control such as AU-Rich Element (ARE) and lipoxygenase differentiation control elements. Bovine FSH(beta) 3ʹUTR down-regulated reporter expression in (alpha)T3-1 and NIH3T3 cells, but not in HEK 293 cells, suggesting the involvement of a cell-specific factor or mechanism. The presence of a 3ʹUTR did not influence reporter mRNA stability, but it did decrease its association with polysomes, indicating that the downregulatory effect may be exerted at the translational level. The segment spanning 601–800 bases (U4) of the bovine FSH(beta) 3ʹUTR was found to be the most effective downregulating segment, its effect being equal to that of the full-length 3ʹUTR. RNA electrophoretic mobility shift assay with U4 showed the presence of specific transfactors in the cytosolic preparations of bovine pituitary and the cell lines. U4 contained an ARE that appeared to be functional, because the mutated U4 ARE was ineffective in downregulating the reporter expression and inhibiting [32P]-labeled U4-transfactor complex formation. Downregulation of reporter activity by the full-length 3ʹUTR and U4 could be overcome by overexpression of HuR, a protein known to stabilize ARE-containing mRNAs in NIH3T3 cells, but not in the (alpha)T3-1 cells, once again indicating that factors other than HuR may also be involved in the regulation of FSH(beta) in the pituitary.