Deciphering p53-regulated alternative transcription and splicing

• Applicants: Privatdozent Dr. Martin Fischer; Professor Dr. Steve Hoffmann
• Subject Area: General Genetics and Functional Genome Biology ; Bioinformatics and Theoretical Biology ; Cell Biology
• Term: since 2021
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 460757154

The determination of mechanisms by which genetic variation produces phenotypic differences affecting tumor development is a primary goal of cancer research. Efforts in this respect have previously focused on mRNA levels altered through epigenetic and transcriptional regulation. Recent studies, however, show that altering isoform abundance is also a common phenomenon in cancer development. The majority of human genes give rise to multiple transcripts that may code for proteins with diverse or even antagonistic biological functions. Importantly, the two major mechanisms that produce this isoform diversity, ie. alternative splicing and alternative transcription via auxiliary transcription start sites (TSS) are regulated by transcription factors (TF). For instance, it has been shown that splicing may depend on transcription factor-driven promoter activity, which is coupled to the elongation rate. In addition, transcription factors are able to alter the genomic splicing landscape by controlling the expression of splicing regulators. Via binding to alternate promoters or enhancers, transcription factors may induce alternative transcription even more directly. Notably, DNA methylation appears to influence alternative promoter usage. Although it is known that transcription factors can influence the isoform landscape in a given cell, the detailed underlying molecular mechanisms are largely unresolved. The p53 signaling pathway is one of the most prominent examples of dysregulation in human cancer, but the consequences of p53 signaling on mRNA isoform abundance are largely unexplored. The overall objective of this research program is to establish a series of protocols and methods to analyze the influence of the transcription factor (TF) p53 on the epigenomic and transcriptomic landscape. Here, we specifically focus on the hypothesis that the transcription factor p53 regulates alternative splicing and may lead to the use of alternative transcription start sites of direct or indirect target genes. The resulting alterations may contribute to p53’s tumor suppressor function. We propose to combine a number of next-generation sequencing experiments and analysis strategies to assess p53-dependent isoform abundance, p53-dependent transcription initiation, p53-induced epigenomic changes, and p53-dependent alternative splicing. We seek to obtain a deeper understanding of TF-mediated isoforms and their underlying mechanisms and will thereby provide resources to simplify future research endeavors. On a broader scale, the proposed research provides a blueprint for comprehensively investigating the effect of TFs on the epigenomic and transcriptomic level. The bioinformatics solutions developed here to link quantitative and qualitative changes of the transcriptome with changes in thechromatin conformation and DNA methylome will facilitate similar research in the future.

• DFG Programme: Research Grants