Abstract
Cytokinesis involves temporally and spatially coordinated action of the cell cycle and cytoskeletal and membrane systems to achieve separation of daughter cells. To dissect cytokinesis mechanisms it would be useful to have a complete catalog of the proteins involved, and small molecule tools for specifically inhibiting them with tight temporal control. Finding active small molecules by cell-based screening entails the difficult step of identifying their targets. We performed parallel chemical genetic and genome-wide RNA interference screens in Drosophila cells, identifying 50 small molecule inhibitors of cytokinesis and 214 genes important for cytokinesis, including a new protein in the Aurora B pathway (Borr). By comparing small molecule and RNAi phenotypes, we identified a small molecule that inhibits the Aurora B kinase pathway. Our protein list provides a starting point for systematic dissection of cytokinesis, a direction that will be greatly facilitated by also having diverse small molecule inhibitors, which we have identified. Dissection of the Aurora B pathway, where we found a new gene and a specific small molecule inhibitor, should benefit particularly. Our study shows that parallel RNA interference and small molecule screening is a generally useful approach to identifying active small molecules and their target pathways.
Highlights
Small molecule inhibitors are useful tools for studying dynamic biological processes
Parallel Screening Protocols To identify all genes required for cytokinesis, and small molecules that target their products, we developed an assay for both comprehensive functional genomic and large-scale chemical genetic screens in cultured Drosophila cells
Small Molecule Screening Results From approximately 51,000 small molecules that included a mixture of commercial ‘‘drug-like’’ molecules, natural product extracts, and natural-product-like libraries synthesized at the Institute of Chemistry and Cell Biology, we identified 50 small molecule inhibitors of cytokinesis, and selected 25 of the most potent and readily available for further analysis (Table S1)
Summary
Small molecule inhibitors are useful tools for studying dynamic biological processes. The targets of small molecules have been identified by methods based on physical affinity, for example, affinity chromatography (Harding et al 1989). These require chemical modification of the small molecule and suffer the limitation that irrelevant proteins will bind in addition to the authentic target. A complementary method is to use information on the biological activity of the small molecule to identify the cellular pathway it perturbs. In some cases an educated guess can be successful (Mayer et al 1999), but to be generally useful, the biological activity of a small molecule would need to be systematically compared to the effect of perturbing different cellular pathways. We develop a parallel screening strategy for finding small molecules that inhibit the biological process of cytokinesis, the genes required for this process, and, by cross-comparison of phenotypes, information on the protein targets of the small molecules
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
