Genetic Screen for Regulators of POL4, a DNA Repair Polymerase in Saccharomyces cerevisiae

Author

• Pasang Dolma Sherpa, CUNY Hunter CollegeFollow

Date of Award

Spring 5-1-2026

Document Type

Thesis

Degree Name

Master of Arts (MA)

Department

Biological Sciences

First Advisor

Professor Diego Loayza

Second Advisor

Prof. Hualin Zhong

Academic Program Adviser

Prof. Diego Loayza

Abstract

In Saccharomyces cerevisiae, DNA polymerase 4 (POL4) is the beta repair polymerase, the product of the POL4 gene, and is involved in base excision repair (BER) and microhomology-mediated end joining (MMEJ). Despite its involvement in these repair pathways and its conservation across eukaryotes, deletion of POL4 shows no detectable phenotype under standard laboratory conditions. I hypothesized that unknown genes act as backup systems, providing redundant activities that allow pol4Δ cells to survive. To test this, a genetic screen for mutants showing synthetic lethality with pol4Δ was implemented. I constructed a strain lacking the POL4 gene and carrying a galactose-inducible PGAL-MYC-POL4 construct, enabling conditional expression of Pol4: the protein is produced in galactose medium and absent in dextrose medium. Following ethyl methanesulfonate (EMS) mutagenesis, approximately 14,000 colonies were screened for dextrose-sensitive growth, identifying mutants that require Pol4 for viability. I describe the isolation of one candidate mutant (13-2) which segregates as a single gene in tetrad dissection, displaying 2:2 segregation of the dextrose-sensitive phenotype consistent with a single Mendelian locus. This candidate could represent an activity redundant with Pol4, on which pol4Δ cells rely for viability. This pilot round of screening supports that the conditional system and screening pipeline are working as intended and suitable for continued identification of synthetic lethal interactions with POL4 and pathways that contribute to genome stability.

Recommended Citation

Sherpa, Pasang Dolma, "Genetic Screen for Regulators of POL4, a DNA Repair Polymerase in Saccharomyces cerevisiae" (2026). CUNY Academic Works.
https://academicworks.cuny.edu/hc_sas_etds/1510