C. elegans THSC/TREX-2 deficiency causes replication stress and genome instability

Transcription is an essential process of DNA metabolism, yet it makes DNA more susceptible to DNA damage. THSC/TREX-2 is a conserved eukaryotic protein complex with a key role in mRNP biogenesis and maturation that prevents genome instability. One source of such instability is linked to transcription as shown in yeast and human cells, but the underlying mechanism and whether is universal is still unclear. To get further insight in the putative role of THSC/TREX-2 in genome integrity we have used Caenorhabditis elegans mutants of the THP-1 and DSS-1 members of THSC/TREX-2. These mutants show similar defective meiosis, DNA damage accumulation and activation of the DNA damage checkpoint. However, they differ regarding replication defects as determined by dUTP incorporation in the germline. Interestingly, this specific thp-1 phenotype can be partially rescued by overexpression of RNase H. Furthermore, both mutants show a mild increase in the H3S10P mark previously shown to be linked to DNA-RNA hybrid-mediated genome instability. These data support the view that both THSC/TREX-2 factors prevent transcription-associated DNA damage derived from DNA-RNA hybrid accumulation by separate means.

Nrd1p identifies aberrant and natural exosomal target messages during the nuclear mRNA surveillance in Saccharomyces cerevisiae

In all eukaryotes, selective nuclear degradation of aberrant mRNAs by nuclear exosome and its cofactors TRAMP, and CTEXT contribute to the fidelity of the gene expression pipeline. In the model eukaryote, Saccharomyces cerevisiae, the Nrd1p-Nab3p-Sen1p (NNS) complex, previously known to be involved in the transcription termination and matured 3’-end formation of vast majority of non-coding and several coding RNAs, is demonstrated to universally participate in the nuclear decay of various kinds of faulty messages in this study. Consistently, nrd1-1/nrd1-2 mutant cells display impairment of the decay of all kinds of aberrant mRNAs, like the yeast mutants deficient in Rrp41p, Rrp6p, and Rrp4p. nrd1ΔCID mutation (consisting of Nrd1p lacking its CID domain thereby abrogating its interaction with RNAPII) however, abolishes the decay of aberrant messages generated during early phases of mRNP biogenesis (transcription elongation, splicing and 3’-end maturation) without affecting the decay rate of the export-defective mRNAs. Mutation in the 3’-end processing factor, Pcf11p, in contrast, displayed a selective abolition of the decay of the aberrant mRNAs, generated at the late phase of mRNP biogenesis (export-defective mRNAs) without influencing the faulty messages spawned in the early phase of mRNP biogenesis. Co-transcriptional recruitment of Nrd1p onto the faulty messages, which relies on RNAPII during transcription elongation and on Pcf11p post transcription, is vital for the exosomal decay of aberrant mRNAs, as Nrd1p deposition on the export-defective messages led to the Rrp6p recruitment and eventually, their decay. Thus, presence of the ‘Nrd1p mark’ on aberrant mRNAs appears rate-limiting for the distinction of the aberrant messages from their normal functional counterparts.Author’s SummaryAberrant/faulty mRNAs generated from the deficiencies in any of the mRNP biogenesis events are promptly eliminated by the nuclear exosome and its cofactors TRAMP and CTEXT complexes. These machineries work relentlessly in the nucleus to detect all kinds of aberrant mRNAs and selectively target them for destruction. However, initial detection of a minuscule amount of aberrant mRNA in the vast background of normal mRNAs is quite challenging and its mechanism remains elusive. In this work, we demonstrate that, the trimeric Nrd1p-Nab3p-Sen1p complex, previously implicated in the transcription termination of diverse non-coding RNAs and a handful of mRNAs, constitute an integral component of the nuclear mRNA surveillance mechanism in baker’s yeast Saccharomyces cerevisiae. Major component of this complex, Nrd1p is demonstrated to be recruited selectively onto various classes of representative model aberrant messages either co-transcriptionally by RNA Polymerase II or post-transcriptionally by Pcf11p. Binding of Nrd1p to the export-defective special mRNAs further leads to the recruitment of Rrp6p on to them thereby leading to their degradation. NNS complex thus plays a vital role of initially recognizing the faulty messages and further assists in the recruitment of the nuclear exosome for their prompt elimination.