An unknown essential function of tRNA splicing endonuclease is linked to the integrated stress response and intron debranching

• Published in: Genetics (Austin) 2023-05, Vol.224 (2)
• Main Authors: Hurtig, Jennifer E, van Hoof, Ambro
• Format: Article
• Language: English
• Subjects: Amino acids; Biodegradation; Degradation; Endonuclease; Fungal Genetics and Genomics; Genetics; Hypoplasia; Introns; Mutants; Mutation; RNA Precursors - genetics; RNA Splicing; RNA, Transfer - genetics; Saccharomyces cerevisiae - metabolism; Splicing; Substrates; Suppressors; Transcriptomes; Transfer RNA; tRNA; Yeast; Yeasts
• ISSN: 1943-2631; 0016-6731; 1943-2631
• DOI: 10.1093/genetics/iyad044

Abstract tRNA splicing endonuclease (TSEN) has a well-characterized role in transfer RNA (tRNA) splicing but also other functions. For yeast TSEN, these other functions include degradation of a subset of mRNAs that encode mitochondrial proteins and an unknown essential function. In this study, we use yeast genetics to characterize the unknown tRNA-independent function(s) of TSEN. Using a high-copy suppressor screen, we found that sen2 mutants can be suppressed by overexpression of SEN54. This effect was seen both for tRNA-dependent and tRNA-independent functions indicating that SEN54 is a general suppressor of sen2, likely through structural stabilization. A spontaneous suppressor screen identified mutations in the intron-debranching enzyme, Dbr1, as tRNA splicing-independent suppressors. Transcriptome analysis showed that sen2 mutation activates the Gcn4 stress response. These Gcn4 target transcripts decreased considerably in the sen2 dbr1 double mutant. We propose that Dbr1 and TSEN may compete for a shared substrate, which TSEN normally processes into an essential RNA, while Dbr1 initiates its degradation. These data provide further insight into the essential function(s) of TSEN. Importantly, single amino acid mutations in TSEN cause the generally fatal neuronal disease pontocerebellar hypoplasia (PCH). The mechanism by which defects in TSEN cause this disease is unknown, and our results reveal new possible mechanisms. Single amino acid mutations in tRNA-splicing endonuclease (TSEN) cause the generally fatal neuronal disease pontocerebellar hypoplasia. TSEN has long been characterized as an enzyme important for making tRNAs; however, its other functions have only been partially been determined and may be relevant to the disease. Here, Hurtig and van Hoof use a yeast genetic approach to characterize additional functions of TSEN, providing new insight into the physiological consequences of its mutation, including an unanticipated activation of the integrated stress response.