Mechanisms and consequences of functional divergence in transcription factor paralogs

• Applicant: Professor Dr. Felix Jonas
• Subject Area: General Genetics and Functional Genome Biology ; Biochemistry ; Bioinformatics and Theoretical Biology
• Term: since 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 542757098

Transcriptional gene regulation is a fundamental ability of living organisms and relies on transcription factors (TF) and the regulatory DNA elements they bind to. By binding to multiple regulatory sites and recruiting effectors to activate or repress the associated genes, TFs form the hubs of the gene regulatory network (GRN). Due to this central position, the emergence of new TFs – predominantly through gene duplication and divergence – is linked to evolutionary breakthroughs, such as multicellularity. However, compared to their present-day functions, the molecular mechanisms generating these functions remain elusive. Most previous research into TF duplicates (paralogs) evolution was limited to individual examples and lacked a unifying framework. In this project, we thus seek a general understanding of the molecular mechanisms that drive the functional divergence of TF duplicates and connect the initial sequence changes to the adaptive rewiring of the GRN. Based on our previous study on the evolution of binding specificity, we propose that, after TF duplication, amino acid (AA) substitutions in intrinsically disordered regions (IDRs) rearrange protein-protein interactions (PPI) changing TF target genes, upstream regulators, and downstream effectors to cumulatively rewire the GRN. To validate this model, we will examine 30 structurally and functionally diverse S. cerevisiae TF paralog pairs through four complementary hypothesis addressing distinct aspects: 1. Gene duplication enables TFs to explore completely new functions. 2. Emerging interactions with other TFs diversify binding specificity. 3. Complementing binding divergence, paralogs can also diverge through effectors. 4. Substitutions in the IDR cumulatively drive divergence between paralogs. To this end, we will compare target genes of TF paralogs across yeast species, explore the difference in the PPI network between paralogs, and finally dissect the AA substitutions distinguishing them. In practice, we will combine CRISPR-Cas9 gene-editing with cutting edge profiling techniques for genomic binding (ChEC-seq) and proteome-wide PPIs (BioID). Furthermore, we will leverage bioinformatics and computational data analysis to explore the expected genome- / proteome-wide datasets and integrate the existing knowledge and sequences information of S. cerevisiae. For each paralog pair, we will describe PPI differences and examine their effect on target selection, explore the adaptability of their binding profiles along evolution, and, for representative pairs, identify divergence-causing AA substitutions. Together, our comprehensive analysis across the full TF spectrum will delineate the molecular mechanisms governing TF evolution and its consequences for GRN, and which probably also acted on the structurally related TFs of more complex, multicellular species, that equally arose from gene duplications.

• DFG Programme: Research Grants