Molecular analysis of translational regulation of thyroid hormone receptor alpha mRNA

Abstract

Amphibian metamorphosis proceeds under the control of thyroid hormone (TH) and thyroid hormone receptor (TR). TRs are encoded by two genes, TRα and TRβ. TRβ mRNA and protein rise along with the increase in endogenous TH. On the other hand TRα mRNA increases during metamorphosis, whereas TRα protein remains constant, suggesting that post‐transcriptional events control the levels of TRα protein. But the regulation of TRα protein expression remains largely unknown. We focused on the region of the TRα 5′UTR. At first, we determined nucleotide sequences of the TRα 5′UTR in amphibians by 5′‐RACE. There are highly conserved reigions in the TRα 5′UTR among the amphibians. We found that the TRα 5′UTR significantly repressed TRα mRNA translation as a cis‐regulatory element in Xenopus laevis A6 (XLA6) cells. Moreover, the TRα 5′UTR suppressed of a downstream open reading frame translation in both reticulocyte lysate and wheat germ extract. Successive deletions of the TRα 5′UTR revealed five elements to repress downstream translation. We analyzed two of five elements by extensive point mutations. One of elements is the GC‐rich region and the other is an upstream open reading frame (uORF). Most single base mutations in the GC‐rich region severely eliminated the repression of translation, while base substitutions didn't affect translational regulation by uORF.