Inclusion Biogenesis and Reactivation of Persistent Chlamydia trachomatis Requires Host Cell Sphingolipid Biosynthesis

Abstract

Chlamydiae are obligate intracellular pathogens that must coordinate the acquisition of host cell-derived biosynthetic constituents essential for bacterial survival. Purified chlamydiae contain several lipids that are typically found in eukaryotes, implying the translocation of host cell lipids to the chlamydial vacuole. Acquisition and incorporation of sphingomyelin occurs subsequent to transport from Golgi-derived exocytic vesicles, with possible intermediate transport through endosomal multivesicular bodies. Eukaryotic host cell-derived sphingomyelin is essential for intracellular growth of Chlamydia trachomatis, but the precise role of this lipid in development has not been delineated. The present study identifies specific phenotypic effects on inclusion membrane biogenesis and stability consequent to conditions of sphingomyelin deficiency. Culturing infected cells in the presence of inhibitors of serine palmitoyltransferase, the first enzyme in the biosynthetic pathway of host cell sphingomyelin, resulted in loss of inclusion membrane integrity with subsequent disruption in normal chlamydial inclusion development. Surprisingly, this was accompanied by premature redifferentiation to and release of infectious elementary bodies. Homotypic fusion of inclusions was also disrupted under conditions of sphingolipid deficiency. In addition, host cell sphingomyelin synthesis was essential for inclusion membrane stability and expansion that is vital to reactivation of persistent chlamydial infection. The present study implicates both the Golgi apparatus and multivesicular bodies as key sources of host-derived lipids, with multivesicular bodies being essential for normal inclusion development and reactivation of persistent C. trachomatis infection.

Author Summary

The genus Chlamydia is composed of a group of obligate intracellular bacterial pathogens that cause several human diseases of medical significance. C. trachomatis is the most commonly encountered sexually transmitted pathogen, as well as the leading cause of preventable blindness worldwide. The prevalence of chlamydial infections, and the extraordinary morbidity and health care costs associated with chronic persisting disease, justifies the research efforts in this area of microbial pathogenesis. Despite their clinical importance, the mechanisms by which these intracellular bacteria obtain nutrients essential to their growth remain enigmatic. Acquisition of sphingolipids, from the cells that chlamydiae infect, is essential for bacterial propagation. This study identifies a requirement for the lipid sphingomyelin from the infected host cell for bacterial replication during infection, and for long-term subsistence in persistent chlamydial infection. Blockage of sphingomyelin acquisition results in premature release of bacteria, a reduced bacterial number, and failure of the bacteria to cause a persisting infection. In this study, we have identified and subsequently disrupted specific sphingomyelin transport pathways, providing important implications on therapeutic intervention targeting this successful microbial pathogen.