The Effect of tRNA([Ser]Sec) Isopentenylation on Selenoprotein Expression

Transfer RNA([Ser]Sec) carries multiple post-transcriptional modifications. The A37G mutation in tRNA([Ser]Sec) abrogates isopentenylation of base 37 and has a profound effect on selenoprotein expression in mice. Patients with a homozygous pathogenic p.R323Q variant in tRNA-isopentenyl-transferase (TRIT1) show a severe neurological disorder, and hence we wondered whether selenoprotein expression was impaired. Patient fibroblasts with the homozygous p.R323Q variant did not show a general decrease in selenoprotein expression. However, recombinant human TRIT1(R323Q) had significantly diminished activities towards several tRNA substrates in vitro. We thus engineered mice conditionally deficient in Trit1 in hepatocytes and neurons. Mass-spectrometry revealed that hypermodification of U(34) to mcm(5)Um occurs independently of isopentenylation of A(37) in tRNA([Ser]Sec). Western blotting and (75)Se metabolic labeling showed only moderate effects on selenoprotein levels and (75)Se incorporation. A detailed analysis of Trit1-deficient liver using ribosomal profiling demonstrated that UGA/Sec re-coding was moderately affected in Selenop, Txnrd1, and Sephs2, but not in Gpx1. 2′O-methylation of U(34) in tRNA([Ser]Sec) depends on FTSJ1, but does not affect UGA/Sec re-coding in selenoprotein translation. Taken together, our results show that a lack of isopentenylation of tRNA([Ser]Sec) affects UGA/Sec read-through but differs from a A37G mutation.