High Throughput Assay for Beta-Catenin
Published 28 August, 2007
Date: August 27, 2007
Project Title: High Throughput Imaging Assay for Beta-Catenin*
Principal Investigator: Patrick M. McDonough, Ph.D.
Abstract: Cancer is one of the most tragic afflictions in modern society, and often results from alterations in the mitotic process. In normal cells, a-catenin is found predominantly associated with the plasma membrane. However, in tumor cells, a-catenin redistributes to the nucleus where it interacts with transcription factors to stimulate the expression of proteins that promote mitosis. The pathway is also important in CNS-related disorders ranging including manic depression, and Alzheimer's. In response to PAR-06-545 "Solicitation of Assays for High Throughput Screening (HTS) in the Molecular Libraries Screening Centers Network (R03)" we are submitting an assay to quantify the cellular distribution of a-catenin. The assay involves exposing HeLa cells to test compounds that are likely to alter a-catenin distribution; the cells are then immunostained for endogenous a- catenin and photographed using high content robotic microscopy workstations. Specialized image analysis software is used to quantify cellular a-catenin localization. In preliminary experiments, an inhibitor of glycogen synthase kinase 3-a (GSK-3), an enzyme typically regulated by the Wnt-signal transduction pathway, induced nuclear localization of a-catenin and this was quantified by the assay with a Z' score of 0.70. The proposed assay will help identify chemicals that are likely to regulate GSK-3 or other proteins that control a-catenin localization. Discovery of such chemicals will contribute to our understanding of the regulation of a-catenin localization and potentially help lead to development of novel therapeutics, depression, and dementia.
Keywords: cadherin, drug discovery /isolation, high throughput technology, molecular /cellular imaging, protein localization, serine threonine protein kinase Wnt gene /protein, antineoplastic, biological signal transduction, chemical registry /resource, image processing, robotics HeLa cell, NIH Roadmap Initiative tag, bioimaging /biomedical imaging, biotechnology.
Media Contact: Valerie M. Hilberg, Direct dial (619) 987-0733
*The project described was supported by Grant Number R03MH082378 from the National Institute on Mental Health.