Identification of Interacting Proteins Via a Yeast Genetic Screen
Location
CSU 253/254/255
Start Date
12-4-2004 1:45 PM
End Date
12-4-2004 3:15 PM
Student's Major
Biological Sciences
Student's College
Science, Engineering and Technology
Mentor's Name
Marilyn Hart
Mentor's Department
Biological Sciences
Mentor's College
Science, Engineering and Technology
Description
Actin, a component of all eukaryotic cells, contributes to cell motility, shape and organization. In striated muscle, actin capping protein (CP) binds to the barbed end of the actin filament at the Zline, directing and maintaining proper organization. CP is a heterodimer composed of alpha and beta subunits. Three isoforms of the beta subunit have been identified: betal (pi), beta! (p2), betaS (p3). The beta isoforms are produced via alternative splicing of one gene and share 90% sequence identity. Previous studies revealed that the beta isoforms have distinct localizations in cardiac myocytes with pi localizing to the Z-line and p2 localizing to the cell periphery and intercalated discs. In transgenic studies, the pi isoform was unable to functionally replace the p2 isoform and vice-versa. These studies suggest that the unique function of the beta isoforms may be due to interactions with additional cellular components. The purpose of this research is to identify proteins that interact with the pi and p2 isoforms of actin CP using a yeast two-hybrid genetic screen. The genetic screen relies upon identifiable gene expression induced by protein interactions. The necessary plasmid constructs have been generated and their orientations confirmed. Western blot analysis revealed that the constructs express the pi and p2 proteins. A small scale preliminary screen is underway.
Identification of Interacting Proteins Via a Yeast Genetic Screen
CSU 253/254/255
Actin, a component of all eukaryotic cells, contributes to cell motility, shape and organization. In striated muscle, actin capping protein (CP) binds to the barbed end of the actin filament at the Zline, directing and maintaining proper organization. CP is a heterodimer composed of alpha and beta subunits. Three isoforms of the beta subunit have been identified: betal (pi), beta! (p2), betaS (p3). The beta isoforms are produced via alternative splicing of one gene and share 90% sequence identity. Previous studies revealed that the beta isoforms have distinct localizations in cardiac myocytes with pi localizing to the Z-line and p2 localizing to the cell periphery and intercalated discs. In transgenic studies, the pi isoform was unable to functionally replace the p2 isoform and vice-versa. These studies suggest that the unique function of the beta isoforms may be due to interactions with additional cellular components. The purpose of this research is to identify proteins that interact with the pi and p2 isoforms of actin CP using a yeast two-hybrid genetic screen. The genetic screen relies upon identifiable gene expression induced by protein interactions. The necessary plasmid constructs have been generated and their orientations confirmed. Western blot analysis revealed that the constructs express the pi and p2 proteins. A small scale preliminary screen is underway.
