Stress adaptation and regulation of gene expression in eukaryotic cells via alternative transcriptional and translational mechanisms
Project is carried out within the TEAM programme of the Foundation for Polish Science, 4.4 SG OP
Global gene expression is subject to systematic reprogramming during cell cycle, development, metabolic changes, stress response and programmed cell death. These transitions, mediated at the transcriptional and postranscriptional levels, involve a massive flux of cellular RNAs and proteins.
The major aim of this project is to establish connections between regulatory mechanisms that that use non-coding and coding properties of RNA molecules and orchestrate changes in gene expression programs during the response to physiological and stress-induced stimuli in eukaryotic cells. We will use a model organism, yeast Saccharomyces cerevisiae, where several of these processes have been characterized to some extent and are relatively straightforward to follow, but their mutual relationships are still not fully verified. We will focus on some specific and less well-recognized elements of the regulatory network that execute programmed changes that accompany cellular response to chosen stress conditions and are controlled at the level of transcription and translation, often through different classes of noncoding RNAs.
Biogenesis of non-coding RNAs
Our recent work on transcription termination and processing of snoRNAs revealed the mechanisms by which the same RNA polymerase II produces different RNA molecules, polyadenylated mRNAs and snoRNAs lacking poly(A) tails. We confirmed that termination of snoRNAs occurs predominantly at the specific Nrd1/Nab3-dependent site, while the second region acts as a fail-safe terminator. We showed that termination at both sites is followed by precursor polyadenylation by a major poly(A) polymerase Pap1 with the contribution by the alternative poly(A) polymerase Trf4.
Functional diversity of non-coding RNAs
The project “Functional diversity of non-coding RNAs” realized by the NOCORE (Non-Coding RNA Enterprise) consortium aims at furthering our understanding of long non-coding RNAs (lncRNA) function in yeast and higher eukaryotic cells. Some of these ncRNAs are abundantly expressed in cancer cells and were proposed to have regulatory roles in signaling pathways. We want to unravel the full potential of natural lncRNAs, as well as those derived from housekeeping RNAs (hncRNAs) (rRNAs, tRNAs and other), in regulation of gene expression in Eukaryotes. This major objective encompasses the discovery of novel classes of ncRNAs, understanding the complexity of their synthesis and maturation as well as regulatory mechanisms, and finally deciphering their function by identifying the substrates they target and processes they act.
Transcription termination by RNA polymerase I
Following co-transcriptional endonculeolytic cleavage of the nascent 35S precursor by Rnt1, the remaining 5’ RNA fragment still associated with Pol I recruits 5’-3’ exonuclease Rat1, which then catches and “torpedoes” the polymerase at a downstream pause site resulting in its release from the DNA template. As this mechanism shows striking similarities to the “torpedo” model for Pol II termination on mRNAs, it confirms that two different polymerases utilize parallel mechanisms of transcription termination. Currently we carry on this line, investigating the involvement of the Nrd1/Nab3 complex in PolI termination.
RNA under oxidative stress, apoptosis and autophagy (ribophagy)
Degradation of ribosomal RNA accompanies some stress- and apoptosis- related programmed cell death pathways or adaptive responses. Such behaviour is consistent with a notion that level of ribosomes is monitored to define cell fitness and survival under unfavourable conditions. RNA fragmentation, carried out by a combined action of endo- and exo-nucleases, including Rny1 (ribophagy), Nuc1 and the exosome, is neutralized by cellular defence systems. Similar phenotypes were observed for tRNAs in various organisms, ranging from Tetrahymena and plants to humans. We speculate that accumulation of rRNA degradation intermediates may act as signaling molecules that trigger downstream events leading to adaptation or cell death.