Human RAB14 (Ras-related protein Rab-14) ELISA Kit

Chlamydia trachomatis, an obligate intracellular bacterium, intercepts different trafficking pathways of the host cell to acquire essential lipids for its survival and replication, particularly from the Golgi apparatus via a Rab14-mediated transport. Molecular mechanisms underlying how these bacteria manipulate intracellular transport are a matter of intense study. Here, we show that C. trachomatis utilizes Akt/AS160 signaling pathway to promote sphingolipids delivery to the chlamydial inclusion through Rab14-controlled vesicular transport. C. trachomatis provokes Akt phosphorylation along its entire developmental life cycle and recruits phosphorylated Akt (pAkt) to the inclusion membrane. As a consequence, Akt Substrate of 160 kDa (AS160), also known as TBC1D4, a GTPase Activating Protein (GAP) for Rab14, is phosphorylated and therefore inactivated. Phosphorylated AS160 (pAS160) loses its ability to promote GTP hydrolysis, favoring Rab14 binding to GTP. Akt inhibition by an allosteric isoform-specific Akt inhibitor (iAkt) prevents AS160 phosphorylation and reduces Rab14 recruitment to chlamydial inclusions. iAkt further impairs sphingolipids acquisition by C. trachomatis-inclusion and provokes lipid retention at the Golgi apparatus. Consequently, treatment with iAkt decreases chlamydial inclusion size, bacterial multiplication, and infectivity in a dose-dependent manner. Similar results were found in AS160-depleted cells. By electron microscopy, we observed that iAkt generates abnormal bacterial forms as those reported after sphingolipids deprivation or Rab14 silencing. Taken together, our findings indicate that targeting the Akt/AS160/Rab14 axis could constitute a novel strategy to limit chlamydial infections, mainly for those caused by antibiotic-resistant bacteria.

The pathogenic traits of TlyA proteins of Mycobacterium tuberculosis are not known. Expressions of TlyA in bacteria that do not express endogenous TlyA adhere better to RAW264.7 macrophages and get phagocytosed efficiently. The internalized bacteria avoid acidification to the extent of >65% in the case of both TlyA-expressing E. coli and M. smegmatis. Consistent with this observation, we have observed decreased co-localizaton of Lysosomal Membrane Associated Protein-1 (similar to 35%), Early Endosomal Antigen-1 (similar to 34%), Rab5 (similar to 30%) and Rab7 (similar to 35%) and enhanced co-localizaton of Rab14 (similar to 80%) on both TlyA-expressing bacteria as well as on TlyA-coated latex beads. These results suggest that the mycobacterial TlyA, in general, can modulate phagolysosome maturation pathway immediately after entry into macrophages, while other important molecules may aid the bacterium for long-term, intracellular survival at later point of time.