Quantitative analysis of genetic and environmental factors determining variation in cell growth Naomi Ziv, Mark Siegal and David Gresham Center for Genomics and Systems Biology, New York University Motivation Growth is a fundamental property of cells. Cell growth is defined by three stages: initiation, proliferation and quiescence. Complex networks of genetic and environmental factors regulate each stage. One of the major growth regulators in Saccharomyces cerevisiae is carbon availability. How a cell regulates its rate of growth in response to carbon availability and the extent to which this regulation varies is currently unknown. Using a novel growth phenotyping assay, we are investigating within and between genotype variation in nutrient-regulated growth rate. We aim to identify the genetic determinants that contribute to variation in this response and the role of evolutionary forces in shaping this variation. Experimental method Individual yeast cells are distributed in glass-bottom 96-well plates and imaged every hour. As the cells grow and divide, they form micro-colonies, consisting of the original single cell and its progeny. Custom image analysis algorithms are used to process the images and compute the area of each micro-colony. Growth parameters for ~40,000, individual micro-colonies are calculated by analyzing the change in area over time. Growth rate Lag duration Initial cell size Representative image from a growth phenotyping experiment. Left, unprocessed image, right, processed image fields are imaged in a single experiment One micro-colony at four different time-points Maximal growth rate is calculated as the highest slope of a linear regression that has a R 2 of at least 0.9, Lag duration is defined by the intersection with a horizontal line defined by the initial cell size. We have phenotyped a set of yeast strains: a lab strain, a North America oak strain, a California vineyard strain, the F1 hybrid of an Oak/Vineyard cross and a panel of F2 segregants. (Gerke, et al., 2006). Strains Results Parental and F1 strains were phenotyped at different glucose concentrations. A mixed effect model was fit to identify a significant dependence of growth rate and lag duration on both nutrient concentration, genetic background and their interaction. F2 segregants were phenotyped at a high glucose concentration (4.44 mM). Median segregant growth rates were normalized according to parental phenotypes. As a measure of within genotype phenotypic variance, a mean levene statistic was also calculated and normalized for each segregant. We have identified QTL influencing growth rate variation in this cross using R/qtl (Broman, et al., 2003). References: Broman KW, Wu H, Sen Ś, Churchill GA (2003) R/qtl: QTL mapping in experimental crosses. Bioinformatics 19: Gerke, J, Chen, C, & Cohen, B.(2006) Natural isolates of Saccharomyces cerevisiae display complex genetic variation in sporulation efficiency. GENETICS, 174 (2), Schematic of strains used in this study Future directions We will resolve QTL influencing growth rate and within genotype variability to the causative QTN. We will also phenotype and map growth rate and lag duration (and their variability) at ten-fold lower glucose concentrations. Variation in growth is determined by environmental and genetic factors QTLS influencing between and within genotype variation in growth Phenotypic variation in F2 growth and variance ✖ P: F1: F2: 