Student Theses and Dissertations

Date of Award

2009

Document Type

Thesis

RU Laboratory

Cross Fred Laboratory

Keywords

Saccharomyces cerevisiae, budding yeast cell cycle, G1 cyclins, CLN3, cell type specific regulation

Abstract

Understanding the molecular and biophysical mechanisms that couple the process of cell growth to cell division is one of the major challenges of modern cell biology. Saccharomyces cerevisiae (budding yeast) has been an important model organism to study the coupling between cell growth and cell division. The insights obtained from studies of this unicellular organism have been pivotal for related studies in animal systems. The classical picture that emerged from studies in budding yeast was that cell cycle commitment in G1, at a point called Start, requires growth to a critical cell size. This deterministic model did not address how cell size control can be achieved despite the stochasticity of elementary cellular processes. Furthermore, no clear connection between the commitment at Start and the molecular network controlling the G1/S transition was known. We developed a novel framework for analyzing the precision of cell size control, by combining single-cell time-lapse imaging of fluorescently labeled cells and rigorous mathematical analysis. This allowed us to quantify the contributions of size control and molecular noise to temporal variability of the G1 phase. Comparing wild-type and mutant strains bearing multiple fluorescent cell cycle markers, we found that Start regulatory dynamics can be decomposed into a size sensing module and a completely independent timing module. We identified inactivation of the Whi5 repressor as marking the boundary between the two modules and showed that different G1 cyclins, CLN3 vs. CLN1 and CLN2, control the two modules. We also showed how positive feedback of G1 cyclins CLN1 and CLN2 on their own transcription ensures a fast transition between the two modules and a coherent commitment to cell cycle progression. Difference in cell size at birth is not the only determinant of the differential regulation of Start between mother and daughter cell. Using single-cell analysis, microarrays and chromatin immuno-precipitations we have shown that cell-type specific difference in regulation of Start is also due to regulation of the G1 cyclin CLN3 by daughter-specific transcription factors Ace2 and Ash1. This work demonstrates how asymmetric localization of cell-fate determinants results in cell-type-specific regulation of the cell cycle in budding yeast.

Comments

A thesis presented to the faculty of The Rockefeller University in partial fulfillment of the requirements for the degree of Doctor of Philosophy.

Permanent URL

http://hdl.handle.net/10209/404

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