Using yeast to discover Toxoplasma cell division genes

St. Cloud State University, Saint Cloud MN

Investigators

Abstract

DESCRIPTION (provided by the applicant): The control of cell cycle division patterns that allow apicomplexan parasites to inhabit diverse niches are not understood. In Toxoplasma, for example, division in the definitive host is a endomitotic process called schizogony. Schizogony is followed by a mitotic division in the intermediate host called endodyogeny. How these parasites switch from each division scheme is critically important as the regulation of growth is linked to development and pathogenesis and, therefore, virulence. By employing yeast as a surrogate genetic system, we will discover and clone cell cycle control molecules from the T. gondii. Saccharomyces cerevisiae is a model organism for heterologous complementation of cell cycle mutants and the yeast two-hybrid system. While the Toxoplasma tachyzoite is the most experimentally tractable apicomplexan, the experiments proposed in the specific aims below are not possible in vivo in the Toxoplasma tachyzoite. Discovery and preliminary characterization of cell cycle control proteins using yeast will advance knowledge in apicomplexan cell cycle biology and provide T. gondii and other apicomplexan investigators new directions for experimentation of the unique features of apicomplexan cell division. Specific aim 1 is to identify T. gondii cell cycle control proteins by yeast two-hybrid system screening. Specific aim 2 is to identify T. gondii cell cycle control proteins by heterologous complementation screening in yeast. We will genetically complement conditional cell cycle mutants of Saccharomyces cerevisiae with a T. gondii cDNA library. Specific aim 3 is the creation of a Toxoplasma cell cycle protein-protein interaction matrix. The yeast two-hybrid system has been used to construct genome wide interaction maps. We will mine the emerging Toxoplasma genome database in conjunction with proteins discovered in aims 1 and 2 to generate a matrix of cell cycle control related proteins of T. gondii and their interactions in the yeast two-hybrid system.