# Genomic Instability and Adaptive Evolution Induced by RFA Insufficiency in Saccharomyces cerevisiae

## TL;DR

This study shows how reduced RFA levels in yeast cause genomic instability and adaptive evolution through specific mutations and repair pathway changes.

## Contribution

The study reveals RFA insufficiency's role in inducing a unique mutational spectrum and adaptive MMR gene inactivation in yeast.

## Key findings

- RFA deficiency causes monosomy and terminal deletions, indicating replication stress.
- Loss of heterozygosity is enriched at centromeres and high-GC regions.
- MMR gene inactivation helps yeast survive RFA stress and recover growth.

## Abstract

This study systematically investigated the genomic alterations in Saccharomyces cerevisiae driven by Replication Factor A (RFA) dosage insufficiency using a promoter-replacement strategy combined with mutation accumulation and whole-genome sequencing. Our findings reveal that transcriptional suppression of RFA2 or RFA3 leads to severe growth inhibition. RFA deficiency induces a distinct mutational spectrum characterized by a high frequency of monosomy and terminal deletions, indicative of severe replication stress. Furthermore, loss of heterozygosity is significantly enriched at centromeres and high-GC regions, underscoring the role of RFA in stabilizing intrinsic genomic barriers. Utilizing an APOBEC3B-induced mutagenesis assay, we demonstrate that RFA insufficiency leads to the extensive accumulation of exposed ssDNA with a distinct bias towards the lagging strand template. Notably, we observed that cells spontaneously inactivate Mismatch Repair (MMR) genes, such as MSH2 and PMS1, to survive RFA-induced stress. This hypermutant phenotype grants a certain degree of growth recovery on Low Galactose (LG) medium. Overall, these findings demonstrate that RFA dosage is a key determinant of genomic integrity and elucidate how repair pathway modulation drives adaptive evolution under replication stress.