FtsZ is a guanosine triphosphatase (GTPase) that mediates cytokinesis in bacteria. FtsZ is homologous in structure to eukaryotic tubulin and polymerizes in a similar head-to-tail fashion. The study of tubulin’s function in eukaryotic cells has benefited greatly from specific and potent small molecule inhibitors, including colchicine and taxol. Although many small molecule inhibitors of FtsZ have been reported, none has emerged as a generally useful probe for modulating bacterial cell division. With the goal of establishing a useful and reliable small molecule inhibitor of FtsZ, a broad biochemical cross-comparison of reported FtsZ inhibitors was undertaken. Several of these molecules, including phenolic natural products, are unselective inhibitors that seem to derive their activity from the formation of microscopic colloids or aggregates. Other compounds, including the natural product viriditoxin and the drug development candidate PC190723, exhibit no inhibition of GTPase activity using protocols in this work or under published conditions. Of the compounds studied, only zantrin Z3 exhibits good levels of inhibition, maintains activity under conditions that disrupt small molecule aggregates, and provides a platform for exploration of structure-activity relationships (SAR). Preliminary SAR studies have identified slight modifications to the two sidechains of this structure that modulate the inhibitory activity of zantrin Z3. Collectively these studies will help focus future investigations toward the establishment of probes for FtsZ that fill the roles of colchicine and taxol in studies of tubulin.

These results provide a new model for USP7 deubiquitinase activity through recruitment to oncogenic chromatin loci and regulation of both oncogenic transcription factors and chromatin marks to promote leukemia. Our studies also show that targeting USP7 inhibition could be a therapeutic strategy in aggressive leukemia.

FtsZ is a homolog of eukaryotic tubulin that is widely conserved among bacteria and coordinates the assembly of the cell division machinery. FtsZ plays a central role in cell replication and is a target of interest for antibiotic development. Several FtsZ inhibitors have been reported. We characterized the mechanism of these compounds in bacteria and found that many of them disrupt the localization of membrane-associated proteins, including FtsZ, by reducing the transmembrane potential or perturbing membrane permeability. We tested whether the reported phenotypes of a broad collection of FtsZ inhibitors disrupt the transmembrane potential in Bacillus subtilis strain 168. Using a combination of flow cytometry and microscopy, we found that zantrin Z1, cinnamaldehyde, totarol, sanguinarine, and viriditoxin decreased the B. subtilis transmembrane potential or perturbed membrane permeability, and influenced the localization of the membrane-associated, division protein MinD. These studies demonstrate that small molecules that disrupt membrane function in bacterial cells produce phenotypes that are similar to the inhibition of proteins associated with membranes in vivo, including bacterial cytoskeleton homologs, such as FtsZ. The results provide a new dimension for consideration in the design and testing of inhibitors of bacterial targets that are membrane-associated and provide additional insight into the structural characteristics of antibiotics that disrupt the membrane.

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