Parallel Chemical Genetic and Genome-Wide RNAi Screens Identify Cytokinesis Inhibitors and Targets

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

A palmitoyl transferase chemical genetic system to map ZDHHC-specific S-acylation

The 23 human ZDHHC S-acyltransferases catalyze long-chain S-acylation at cysteine residues across an extensive network of hundreds of proteins important for normal physiology or dysregulated in disease. Here we present a technology platform to directly map the protein substrates of a specific ZDHHC for the first time at the whole proteome level, in intact cells. Structure-guided engineering of paired ZDHHC ‘hole’ mutants and ‘bumped’ chemically tagged fatty acid probes enabled probe transfer to specific protein substrates with excellent selectivity over wild type ZDHHCs. Chemical genetic systems were exemplified for five ZDHHCs (3, 7, 11, 15 and 20), and applied to generate the first de novo ZDHHC substrate profiles, identifying >300 unique and shared substrates across multiple cell lines and Sacylation sites for novel functionally diverse substrates. We expect that this powerful and versatile platform will open a new window on S-acylation biology for a wide range of models and organisms

The Chromosomal Passenger Complex Activates Polo Kinase at Centromeres

INCENP acts as a protein scaffold that integrates the functions of two crucial mitotic kinases, Aurora B and Polo, at centromeres of mitotic chromosomes

The Chromosomal Passenger Complex Activates Polo Kinase at Centromeres

INCENP acts as a protein scaffold that integrates the functions of two crucial mitotic kinases, Aurora B and Polo, at centromeres of mitotic chromosomes

Non-traditional roles of G protein-coupled receptors in basic cell biology

G protein-coupled receptors (GPCR) are key signaling proteins that regulate how cells interact with their environments. Traditional signaling cascades involving GPCRs have been well described and are well established and very important clinical targets. With the development of more recent technologies, hints about the involvement of GPCRs in fundamental cell biological processes are beginning to emerge. In this review, we give a basic introduction to GPCR signaling and highlight some less well described roles of GPCRs, including in cell division and membrane trafficking, which may occur through canonical and non-canonical signaling pathways

Capping protein regulates actin dynamics during cytokinetic midbody maturation

Significance Actin dynamics drive many steps of cell division. Here, we show that the actin capping protein (CP) is unexpectedly involved in midbody maturation, a poorly understood phase of the cell cycle where cells remodel their intercellular bridges to prepare for separation. The loss of CP results in excessive filamentous actin throughout the cell cycle, but only postfurrowing cytokinesis is inhibited. We propose that optimal actin filament function is achieved by a balance between CP-dependent filament capping and formin-driven polymerization. This raises the intriguing possibility that cells utilize specific types of actin filament networks to progress through division. This finding has profound implications on understanding actin-dependent processes such as cell division, migration, adhesion, and morphogenesis.</jats:p