An intact unfolded protein response in Trpt1 knockout mice reveals phylogenic divergence in pathways for RNA ligation

Abstract

Unconventional mRNA splicing by an endoplasmic reticulum stress-inducible endoribonuclease, IRE1, is conserved in all known eukaryotes. It controls the expression of a transcription factor, Hac1p/XBP-1, that regulates gene expression in the unfolded protein response. In yeast, the RNA fragments generated by Ire1p are ligated by tRNA ligase (Trl1p) in a process that leaves a 2′-PO₄ ²⁻ at the splice junction, which is subsequently removed by an essential 2′-phosphotransferase, Tpt1p. However, animals, unlike yeast, have two RNA ligation/repair pathways that could potentially rejoin the cleaved Xbp-1 mRNA fragments. We report that inactivation of the Trpt1 gene, encoding the only known mammalian homolog of Tpt1p, eliminates all detectable 2′-phosphotransferase activity from cultured mouse cells but has no measurable effect on spliced Xbp-1 translation. Furthermore, the relative translation rates of tyrosine-rich proteins is unaffected by the Trpt1 genotype, suggesting that the pool of (normally spliced) tRNA^Tyr is fully functional in the Trpt1−/− mouse cells. These observations argue against the presence of a 2′-PO₄ ²⁻ at the splice junction of ligated RNA molecules in Trpt1−/− cells, and suggest that Xbp-1 and tRNA ligation proceed by distinct pathways in yeast and mammals.

Keywords

• tRNA splicing
• RNA ligase, 2′-phosphotransferase
• translation
• protein folding; mouse genetics