An Estradiol-Inducible Promoter Enables Fast, Graduated Control of Gene Expression in Fission Yeast [preprint]

The fission yeast Schizosaccharomyces pombe lacks a diverse toolkit of inducible promoters for experimental manipulation. Available inducible promoters suffer from slow induction kinetics, limited control of expression levels and/or a requirement for defined growth medium. In particular, no S. pombe inducible promoter systems exhibit a linear dose response, which would allow expression to be tuned to specific levels. We have adapted a fast, orthogonal promoter system with a large dynamic range and a linear dose response, based on β-estradiol-regulated function of the human estrogen receptor, for use in S. pombe. We show that this promoter system, termed Z3EV, turns on quickly, can reach a maximal induction of 20 fold, and exhibits a linear dose response over its entire induction range, with few off target effects. We demonstrate the utility of this system by regulating the mitotic inhibitor Wee1 to create a strain in which cell size is regulated by β-estradiol concentration. This promoter system will be of great utility for experimentally regulating gene expression in fission yeast

Size-Dependent Expression of the Mitotic Activator Cdc25 as a Mechanism of Size Control in Fission Yeast [preprint]

Proper cell size is essential for cellular function (Hall et al., 2004). Nonetheless, despite more than 100 years of work on the subject, the mechanisms that maintain cell size homeostasis are largely mysterious (Marshall et al., 2012). Cells in growing populations maintain cell size within a narrow range by coordinating growth and division. Bacterial and eukaryotic cells both demonstrate homeostatic size control, which maintains population-level variation in cell size within a certain range, and returns the population average to that range if it is perturbed (Marshall et al., 2012; Turner et al., 2012; Amodeo and Skotheim, 2015). Recent work has proposed two different strategies for size control: budding yeast has been proposed to use an inhibitor-dilution strategy to regulate size at the G1/S transition (Schmoller et al., 2015), while bacteria appear to use an adder strategy, in which a fixed amount of growth each generation causes cell size to converge on a stable average, a mechanism also suggested for budding yeast (Campos et al., 2014; Jun and Taheri-Araghi, 2015; Taheri-Araghi et al., 2015; Tanouchi et al., 2015; Soifer et al., 2016). Here we present evidence that cell size in the fission yeast Schizosaccharomyces pombe is regulated by a third strategy: the size dependent expression of the mitotic activator Cdc25. The cdc25 transcript levels are regulated such that smaller cells express less Cdc25 and larger cells express more Cdc25, creating an increasing concentration of Cdc25 as cell grow and providing a mechanism for cell to trigger cell division when they reach a threshold concentration of Cdc25. Since regulation of mitotic entry by Cdc25 is well conserved, this mechanism may provide a wide spread solution to the problem of size control in eukaryotes