REGULATION OF CELL AND ORGAN SIZE IN VIVO

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The eventual size reached by an animal is determined both by the total number of cells as well as the size of individual cells. Total cell number is determined by cell proliferation and cell death. Recent years have witnessed substantial progress our understanding of the regulation of these two processes. In contrast, the mechanisms that regulate the size of individual cells are not well understood. The experiments detailed in this proposal are aimed at understanding the mechanisms that regulate cell size in vivo. A genetic screen using FLP-recombinase induced mitotic recombination in the Drosophila eye was used to identify mutations that resulted in increased cell size. The mutations isolated to date map to six different genetic loci. One of the loci corresponds to the gigas gene which has been characterized by others and results in large polyploid cells. The gigas gene is the Drosophila homologue of the TSC2 gene which is mutated in approximately 70 percent of patients with tuberous sclerosis. One of the loci identified in the screen, rocky has a phenotype that is extremely similar to that of gigas and maps to the location of the TSC1 gene which is mutated in 30 percent of patients with tuberous sclerosis. Another mutation, which increases cell size without altering ploidy, maps to the vicinity of PTEN gene which is mutated in a number of different human cancers. Another, expanded, appears to act in a non-autonomous manner. Two others have not yet been mapped. Specific Aim 1 of this proposal describes the completion of a comprehensive genetic screen to identify mutations that lead to an increase in cell size. Mutations will be sorted into complementation groups and the genes will be mapped. Specific Aim 2 describes a detailed phenotypic analysis of gigas, rocky, wolf and expanded. Experiments will address the precise mechanism by which these mutations increase cell size. In each case the effect of the mutation on ploidy, growth and cycle regulation will be determined and the role of the gene will be studied in the context of known regulators of cell size. Specific Aim 3 describes a general strategy for characterizing the phenotype of the remaining mutations and those that will be identified as the screen moves to completion. An approach to cloning the genes is also described. Analysis of the mutations identified in the screen will contribute to our understanding of cell size regulation in vivo.