A Conserved Stem Loop Motif in the 5′Untranslated Region Regulates Transforming Growth Factor-β1 Translation

Transforming growth factor-beta(1) (TGF-beta(1)) regulates cellular proliferation, differentiation, migration, and survival. The human TGF-beta(1) transcript is inherently poorly translated, and translational activation has been documented in relation to several stimuli. In this paper, we have sought to identify in cis regulatory elements within the TGF-beta(1) 5'Untranslated Region (5'UTR). In silico analysis predicted formation of stable secondary structure in a G/C-rich element between nucleotides + 77 to + 106, and demonstrated that this element is highly conserved across species. Circular dichroism spectroscopy confirmed the presence of secondary structure in this region. The proximal 5'UTR was inhibitory to translation in reporter gene experiments, and mutation of the secondary structure motif increased translational efficiency. Translational regulation of TGF-beta(1) mRNA is linked to altered binding of YB-1 protein to its 5'UTR. Immunoprecipitation-RT-qPCR demonstrated a high basal association of YB-1 with TGF-beta(1) mRNA. However, mutation of the secondary structure motif did not prevent interaction of YB-1 with the 5'UTR, suggesting that YB-1 binds to this region due to its G/C-rich composition, rather than a specific, sequence-dependent, binding site. These data identify a highly conserved element within the TGF-beta(1) 5'UTR that forms stable secondary structure, and is responsible for the inherent low translation efficiency of this cytokine.