Novel molecular mechanisms connect the conserved histone chaperone FACT to chromosome segregation and condensation in yeast and human.

• Applicant: Magdalena Murawska, Ph.D.
• Subject Area: General Genetics and Functional Genome Biology ; Cell Biology
• Term: since 2025
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 553260496

Histone chaperones are a group of proteins that bind histones and guide the assembly and disassembly of nucleosomes which is vital for maintaining genome accessibility and stability. The conserved histone chaperone FACT has been involved in chromatin maintenance during transcription and the silent heterochromatin spreading in fission yeast, S. pombe. Our new unpublished data show that FACT is also involved in mitosis in S. pombe. We have obtained a FACT mutant that exhibits mitotic defects independent of its functions in heterochromatin. Among others, the FACT mutant exhibits: minichromosome loss phenotype, prolonged anaphase, increased occurrence of lagging chromosomes and condensation defects. AlphaFold candidate screen revealed putative novel interaction partners of FACT involved in chromosome segregation. In this research proposal we will study the molecular basis of FACT regulating chromosome segregation in S. pombe and human cells. We will test the hypothesis that FACT regulates chromosome segregation via its novel interactions. We will use life cell imaging, cell cycle ChIPs and state of the art biochemistry approaches to reveal spatiotemporal kinetochore assembly in the wild type and the FACT mutant strains. As a parallel hypothesis, we will test the role of FACT in chromosome condensation in S. pombe. For this purpose, we will apply quantitative chromosome condensation assays and genomic approaches that can precisely measure chromosome folding at the nucleosomal resolution. Further, we will study the conservation of FACT mitotic functions in human cells using knockdowns and pharmacological inhibition of FACT combined with immunofluorescence and life cell imaging. Finally, we will dissect the molecular basis of FACT spatiotemporal dynamics during yeast and human mitosis using ChIP-seq in synchronized cells and life cell imaging. Together, the results of this project will provide important new insights into the molecular functions of FACT in mitosis and their evolutionary conservation, which have so far been little studied. As cell cycle dysregulation is one of the hallmarks of cancer and FACT inactivation has been recently shown to have positive effects in several cancer types, we anticipate that the results of this project will help to understand why cancer cells require FACT for survival and will possibly open up an avenue for the development of new, chaperone-targeting therapies.

• DFG Programme: Research Grants