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Authors: Cortés López, Mariela
Title: Decoding splicing regulatory networks in cancer-relevant genes
Online publication date: 8-Feb-2022
Year of first publication: 2022
Language: english
Abstract: Alternative splicing is a highly regulated cellular process, relevant to the generation of protein diversity. Elements in the sequence of the transcripts regulate splicing in cis. These cis-elements are bound by trans-acting factors which, in their majority, are RNA binding proteins (RBPs). The interactions between cis-elements and trans-acting factors define the splicing code. Meanwhile, errors in splicing regulation have been frequently associated with diseases, highlighting the need to understand the mechanisms and consequences of mis-splicing. The first part of this work describes the development of a high throughput mutagenesis assay that allows us to look at the impact of individual mutations in the splicing of the exon 11 in the proto-oncogene RON. This assay uncovered the splicing network of RON exon 11 and identified HNRNPH as a relevant regulator. We also show how HNRNPH cooperatively regulates RON exon 11 splicing in a switch-like manner. During the second part of this work, I present the story of a spurious isoform, known as ex2Δpart. This isoform is an example of an exonic intron (exitron) with suggested potential as a therapeutic marker. We show how ex2Δpart is instead an artefact of reverse transcription (RT). Using bioinformatical analysis, we describe other artefacts (named “falsitrons”). Our work also suggests new strategies to refine isoform annotation. The previous chapters also serve as a precedent for the second high-throughput study presented here: the mutagenesis of CD19 exon 2. CD19 is the target of the CART(Chimeric Antigen Receptor T)-19 therapy, and mis-splicing of its second exon has been associated with therapy resistance. Here, we present a high-throughput mutagenesis assay that characterises the cis- and trans-regulators in the region between exons 1-3 of CD19. The study also provides new information on the regulatory network of splicing of CART-19 relapse patients. Together, the regulatory networks described in this work contribute to interpreting mutations in two cancer-relevant genes. In addition, this work also presents a collection of distinct approaches to improve splicing annotations. The tools and analysis described here can be extended to other important mis-splicing events, helping to decipher the splicing code.
DDC: 500 Naturwissenschaften
500 Natural sciences and mathematics
Institution: Johannes Gutenberg-Universität Mainz
Department: FB 10 Biologie
Place: Mainz
URN: urn:nbn:de:hebis:77-openscience-f64315fd-cbda-4761-aa85-bcf70c7213ab8
Version: Original work
Publication type: Dissertation
License: CC BY
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Extent: 190 Seiten, Diagramme
Appears in collections:JGU-Publikationen

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