Posttranscriptional regulation of human ADH5/FDH and Myf6 gene expression by upstream AUG codons

Upstream open-reading frames are unusual in mammalian mRNAs. The 5' untrans lated region of ADH5 mRNA contains an upstream open-reading frame (uORF) wi th two possible AUG start codons. Myf6 mRNA contains three tandem AUG repea ts at the translation start site, a rare feature, Mutation at one or both o f the upstream AUG codons in the ADH5 mRNA increased gene expression twofol d in CV-1, NIH/3T3, HeLa, and SL2 cells, Mutation of these AUG codons led t o 3- to 5-fold increases in activity as measured by in vitro translation as says using capped mRNAs. RNA toeprint analysis demonstrated many stalled ri bosomes flanking the AUG codons and secondary structures near the AUGs. Sec ondary structures may increase the ability of ribosomes to recognize the tw o AUGs, despite their poor initiation context. The degree of repression by uAUGs varied significantly depending on the cell lines tested, which may pa rtly explain the differential tissue expression. Myf6 is a critical myogeni c transcription factor with the striking feature of three tandem AUG codons at the translation initiation site. This structure reduced expression; rem oving two of these AUGs led to a doubling of activity in CV-1, HeLa, and NI H/3T3 cells, (C) 2001 Academic Press.