Talk abstracts
Talk on Saturday 09:30-09:45am submitted by Jacob Smith
Mapping the transcriptional effects of the ER stress sensor IRE1 with optogenetic oligomerization and long-read sequencing
Jacob W. Smith (Department of Chemistry and Biochemistry, The Ohio State University), Vladislav Belyy (Department of Chemistry and Biochemistry, The Ohio State University)
Abstract:
The Unfolded Protein Response (UPR) is a eukaryotic stress response pathway that can switch between promoting cellular homeostasis and regulated cell death in response to imbalances in the endoplasmic reticulum (ER). Inositol-requiring enzyme 1 (IRE1) is one of three known ER membrane-resident stress sensors, and its activity in response to stress is thought to promote the cytoprotective aspects of the UPR. IRE1 responds to unfolded proteins in the ER lumen by forming homo-oligomers that enable trans-autophosphorylation and activation of the cytosolic RNase domain, which then facilitates non-canonical splicing of the mRNA of transcription factor XBP1 into an active isoform that induces a sweeping transcriptional program. IRE1 has been shown to play an important role in diseases where cells are faced with chronic ER stress, such as neurodegeneration and diabetes. A lack of direct experimental means of activation has made it difficult to study IRE1 signaling independently of the other UPR sensors, leading us to engineer an IRE1 construct with an added light-inducible oligomerizing domain to cluster and subsequently activate the cytosolic RNase domain. We used long-read sequencing to analyze the transcriptome of human cells expressing this IRE1 construct and subjected to varying exposures of light or chemical stress. Our data have indicated potential new players in the IRE1 signaling pathway and support the hypothesis that IRE1’s RNase domain can degrade a broad set of mRNAs in addition to XBP1, its canonical target. This method has also enabled us to investigate alternative splicing patterns and novel splicing substrates of IRE1. By using a specific and direct method to investigate the role of IRE1 signaling in the complex network of the UPR, our data provide insights into the role of IRE1 in the human cell’s ability to adapt and respond to severe ER stress and may guide the development of future therapeutics.
Keywords: Transcriptomics, Splicing, Stress