Abstract

This thesis addresses the fundamental biological question of how cells achieve different regulations of protein subgroups with cell volume and ploidy (ch. 1). For most proteins, the production is coupled to cell volume, leading to constant concentrations with increasing cell volume. However, strongly DNA-bound proteins, such as histones, are likely needed at constant amounts, which would lead to decreasing concentrations as a function of cell volume. It still remains unclear how cells could achieve to couple the production of some protein subgroups to cell volume, but decouple it from cell volume for other protein subgroups. To answer this question, I focused on the regulation of histone proteins, using the model organism Saccharomyces cerevisiae and made use of genetics in order to achieve a wider range of observable cell volumes (ch. 2.1). Additionally, I performed live-cell fluorescence microscopy (ch. 2.3), reverse-transcription quantitative PCR (ch. 2.4), flow cytometry (ch. 2.5) and single-molecule fluorescence in situ hybridization (ch. 2.6). First, I demonstrate that histone protein and mRNA concentrations decrease with cell volume and are coupled to their respective gene copy number (ch. 3.2 & 3.3). Then, I show that this regulation is achieved by the histone promoter (ch. 3.4), is not an effect of the cell cycle dependent production of histones (ch. 3.7), and most likely does not require direct transcriptional feedback or degradation mechanisms (ch. 3.8). I also detail a minimal mathematical model that describes the dependence of promoter transcription rates with cell volume (ch. 3.5). This model predicts that the behaviour of a histone promoter can be changed to that of most promoters, solely by decreasing its transcription initiation rate. Using a series of promoter truncations, I confirm this theoretical prediction experimentally (ch. 3.6). In summary, this work shows that the regulation of histone proteins with cell volume and genome content is likely an intrinsic property of histone promoters, where direct transcriptional feedback or degradation mechanisms might play a role but are not necessary. The model introduced during this work lays a foundation for a deeper understanding of protein regulation with cell volume and ploidy in general. Interestingly, this regulation is likely conserved across most eukaryotic cells.