Rab6 and Rab11 Regulate Chlamydia trachomatis Development and Golgin-84-Dependent Golgi Fragmentation

Many intracellular pathogens that replicate in special membrane bound compartments exploit cellular trafficking pathways by targeting small GTPases, including Rab proteins. Members of the Chlamydiaceae recruit a subset of Rab proteins to their inclusions, but the significance of these interactions is uncertain. Using RNA interference, we identified Rab6 and Rab11 as important regulators of Chlamydia infections. Depletion of either Rab6 or Rab11, but not the other Rab proteins tested, decreased the formation of infectious particles. We further examined the interplay between these Rab proteins and the Golgi matrix components golgin-84 and p115 with regard to Chlamydia-induced Golgi fragmentation. Silencing of the Rab proteins blocked Chlamydia-induced and golgin-84 knockdown-stimulated Golgi disruption, whereas Golgi fragmentation was unaffected in p115 depleted cells. Interestingly, p115-induced Golgi fragmentation could rescue Chlamydia propagation in Rab6 and Rab11 knockdown cells. Furthermore, transport of nutrients to Chlamydia, as monitored by BODIPY-Ceramide, was inhibited by Rab6 and Rab11 knockdown. Taken together, our results demonstrate that Rab6 and Rab11 are key regulators of Golgi stability and further support the notion that Chlamydia subverts Golgi structure to enhance its intracellular development

Multiple Phosphatidylinositol 3-Kinases Regulate Vaccinia Virus Morphogenesis

Poxvirus morphogenesis is a complex process that involves the successive wrapping of the virus in host cell membranes. We screened by plaque assay a focused library of kinase inhibitors for those that caused a reduction in viral growth and identified several compounds that selectively inhibit phosphatidylinositol 3-kinase (PI3K). Previous studies demonstrated that PI3Ks mediate poxviral entry. Using growth curves and electron microscopy in conjunction with inhibitors, we show that that PI3Ks additionally regulate morphogenesis at two distinct steps: immature to mature virion (IMV) transition, and IMV envelopment to form intracellular enveloped virions (IEV). Cells derived from animals lacking the p85 regulatory subunit of Type I PI3Ks (p85α−/−β−/−) presented phenotypes similar to those observed with PI3K inhibitors. In addition, VV appear to redundantly use PI3Ks, as PI3K inhibitors further reduce plaque size and number in p85α−/−β−/− cells. Together, these data provide evidence for a novel regulatory mechanism for virion morphogenesis involving phosphatidylinositol dynamics and may represent a new therapeutic target to contain poxviruses