PDF, English Restricted access: Repository staff only until 1 September 2026. Login+Download (12MB) | Lizenz: An investigation of poly(C)-binding protein 1 (PCBP1) and its RNA-interactome in cellular stress responses by Gerritsen, Manouk underlies the terms of Creative Commons Attribution-NoDerivatives 4.0

Abstract

RNA-binding proteins (RBPs) play essential roles in regulating gene expression at the posttranscriptional level and are increasingly recognised to be critical determinants of the cellular response to various stresses. Exposure to stress activates a highly conserved pathway called the integrated stress response (ISR) which promotes translational reprogramming and allows cells to adapt to the stress or undergo programmed cell death. It is therefore fundamental to elucidate the dynamic RNA-protein interactions that enable cells to respond to stress. Previous work from our lab revealed that the poly(A) RNA-binding activity of PCBP1 is sensitive to oxidative stress. Specifically, PCBP1 RNA binding activity is reduced upon arsenite-induced oxidative stress in both the nucleus and the cytoplasm in a hepatocellular carcinoma cell line. In addition to its well-established role as an RBP that regulates nearly all steps of mRNA metabolism, PCBP1 has more recently been identified as a cytosolic iron chaperone that facilitates the delivery of iron to ferritin for safe storage and other nonhaem iron-dependent proteins. Hereby, PCBP1 plays an essential role in limiting the toxicity of oxidative free iron ions in cells. Therefore, the RNA-binding and iron-binding functions of PCBP1 might be connected and interlink post-transcriptional regulation of gene expression by oxidative stress and iron overload, acting at the interface of two biochemically interlinked stressors that affect cells. Here, both the effect of RNA on the iron chaperone function of PCBP1 as well as the effect of iron on PCBP1 RNA binding was tested. In addition, CLIP experiments were performed to map the stress-induced changes in the PCBP1 RNA interactome under arsenite-induced oxidative stress and iron overload. Interestingly, the data show that (acute) iron overload did not significantly affect the RNA interactome of PCBP1, suggesting that PCBP1 RNA binding is not significantly affected by iron. Conversely, arseniteinduced oxidative stress demonstrated significant changes in the interaction of PCBP1 with a subset of RNAs. These transcripts were primarily protein-coding and GO enrichment analysis revealed that mRNAs with enhanced PCBP1 binding under stress are involved in RNA metabolism. By contrast, the mRNAs for which PCBP1 binding is reduced upon stress are more related to metabolic enzymes and regulation of cell adhesion and integrins. These observations indicate that the changes in the RNA interactome of PCBP1 upon oxidative stress may contribute to the reprogramming of gene expression during stress, allowing cells to restore cellular homeostasis.