Molecular mechanisms underlying the commitment to division in budding yeast
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Authors
Ghimire, Abhishek
Issue Date
2025-05
Type
Electronic thesis
Thesis
Thesis
Language
en_US
Keywords
Biology
Alternative Title
Abstract
The regulation for cell size homeostasis is evolutionarily conserved and is vital for organismal survival across the tree of life. The coordination between cell growth and division for cell size homeostasis occurs primarily at the G1/S phase transition, termed Start in budding yeast. The transcriptional activation of ≈200 genes in the G1/S regulon by key transcription factor complexes, SBF and MBF, is essential for the progression of cell cycle post Start. This study focuses on the quantitative assessment of the molecular mechanisms of cell size regulation through investigation into the absolute copy number, concentration, localization and coordination of the Start machinery. Our lab has previously shown that the cell size dependent accumulation of SBF and MBF, and their subsequent titration of the target promoters throughout G1 are critical determinant of Start (Dorsey et al. 2018). This work expands on that study by investigating the dependence of Start on the copy number of Swi4, the DNA binding subunit of SBF, under different genetic contexts using single molecule fluctuation microscopy. We found that Swi4 transcription is partly SBF-mediated and is essential for Start transition. Further, a threshold level of about 170 Swi4 molecules titrating SBF binding sites in G1/S promoters predicted the effects of nutrients, ploidy, and G1/S regulatory mutations on cell size. Additionally, this work reveals the expression dynamics of the extremely low abundance and high turnover G1 cyclins, responsible for the phosphorylation of the SBF repressor, Whi5. Building on the past finding in our lab that the G1/S transcription factors assemble in increasing numbers of discrete clusters in G1 (Black et al. 2020), which suggest a possible clustering of their target promoters as well, we provide some preliminary results into the investigation of the super-resolution localization of the G1/S promoters in budding yeast. In combination, our work provides quantitative insights into the mechanisms of cell size regulation in budding yeast.
Description
May2025
School of Science
School of Science
Full Citation
Publisher
Rensselaer Polytechnic Institute, Troy, NY