Our laboratory is actively engaged in characterizing RNA decay pathways within mammalian nuclei. These efforts involve the identification of new pathways and components as well as their substrates. To this end, we use state-of-the-art affinity capture / mass spectrometry approaches and high throughput transcriptome-wide methodologies. However, decay of RNA is by no means just a ‘clean-up-act’ to remove worn out molecules. In addition to its essential role in the quality control of genome expression, RNA turnover is also at the core of gene expression regulation – forming intricate connections to RNA productive systems – thus, being centrally placed to determine RNA/RNP fate. Therefore, we also work towards establishing models for how regulated RNA turnover helps control key biological processes. To this end, we are mainly using mouse embryonic stem cells and their ability to differentiate and elicit various gene expression programs.
Genomes are pervasively transcribed. Our laboratory aims to identify physiologically relevant transcription events from the sea of spurious events that constantly produce unstable transcripts. This includes new aspects of transcription interference and/or activation as well as the putative identification of new physiologically meaningful RNAs. Such uncovering of new gene regulatory concepts created by pervasive transcription is of interest to us in both the ’normal’ situation and in cases where cells are perturbed by diseases such as cancer or neurobiological defects.
Our laboratory has recently taken an interest into the biogenesis, regulation and function of small nucleolar (sno)RNAs. Most vertebrate snoRNAs are encoded within the introns of highly expressed protein-coding or lncRNA genes – so-called snoRNA host genes. The maturation of intron-hosted snoRNAs generally depends on the splicing process, and therefore snoRNA production gives rise to two decoupled, putatively functional RNA species. We are actively investigating the interplay between and regulation of expression of snoRNAs and their hosts at several levels. Examples of some of our published observations within this area of research are: (1) spliced snoRNA host transcripts that reach the cytoplasm are more often subjected to nonsense-mediated mRNA decay compared to general protein-coding genes (Lykke-Andersen, Chen et al., 2014; Lykke-Andersen and Jensen, 2015), (2) nuclear localized spliced snoRNA host transcripts are identified by the PAXT connection and degraded by the RNA exosome (Meola, Domanski et al., 2016; Silla et al., 2018; Wu, Schmid et al., 2020), (3) the excised snoRNA-containing introns are processed into mature snoRNAs via the NEXT complex and the nuclear RNA exosome (Lubas et al., 2015; Giacometti et al., 2017), and, (4) the box C/D subclass of snoRNAs are subjected to an intricate autoregulation involving a cis-acting snoRNA, alternative splicing, snoRNA binding proteins and possibly snoRNA-lncRNA hybrid (Lykke-Andersen, Ardal et al., 2018). In combination, our studies of snoRNA regulation has revealed interesting novel insights into the regulation of eukaryotic gene expression.
In our lab, we use the budding yeast Saccharomyces cerevisiae to study RNA metabolism in the cell nucleus and focus on the relationships between RNA synthesis, decay and export. To identify relevant substrates of key players, we use rapid nuclear protein depletion schemes (‘anchor-away’ or ’degron’ approaches), coupled with genome-wide methodologies for measuring transcription and RNA synthesis as well as RNA fluorescense in-situ hybridization. Nuclear RNA metabolism is not only an intricate component of the gene expression machinery but also provides opportunities for regulation of transcript levels and for establishing cellular RNA homeostasis via functional coupling to transcription and cytoplasmic RNA decay. Identification of such principles is the primary target of our yeast work. Indeed, we have previously identified new paradigms for gene expression control based on polyadenylation, poly(A) binding proteins, nuclear mRNA export and RNA 3’-5’ exonucleolytic decay (Saguez et al. 2008; Schmid et al. 2012; Schmid et al. 2015; Schmid et al. 2018; Tudek et al. 2018). Consequently, our yeast work provides important insights in its own right, but also contributes important inspiration for projects in mammalian cells.