Phthiocerol Dimycocerosates of M. tuberculosis Participate in Macrophage Invasion by Inducing Changes in the Organization of Plasma Membrane Lipids

Catherine Astarie-Dequeker,André Lopez,Christian Chalut,Laurent Le Guyader,Christophe Guilhot,Wladimir Malaga,Fam-Ky Seaphanh,Joanne L Flynn

doi:10.1371/journal.ppat.1000289

Abstract

Phthiocerol dimycocerosates (DIM) are major virulence factors of Mycobacterium tuberculosis (Mtb), in particular during the early step of infection when bacilli encounter their host macrophages. However, their cellular and molecular mechanisms of action remain unknown. Using Mtb mutants deleted for genes involved in DIM biosynthesis, we demonstrated that DIM participate both in the receptor-dependent phagocytosis of Mtb and the prevention of phagosomal acidification. The effects of DIM required a state of the membrane fluidity as demonstrated by experiments conducted with cholesterol-depleting drugs that abolished the differences in phagocytosis efficiency and phagosome acidification observed between wild-type and mutant strains. The insertion of a new cholesterol-pyrene probe in living cells demonstrated that the polarity of the membrane hydrophobic core changed upon contact with Mtb whereas the lateral diffusion of cholesterol was unaffected. This effect was dependent on DIM and was consistent with the effect observed following DIM insertion in model membrane. Therefore, we propose that DIM control the invasion of macrophages by Mtb by targeting lipid organisation in the host membrane, thereby modifying its biophysical properties. The DIM-induced changes in lipid ordering favour the efficiency of receptor-mediated phagocytosis of Mtb and contribute to the control of phagosomal pH driving bacilli in a protective niche.

Highlights

The cell envelope of Mycobacterium tuberculosis (Mtb) plays a major role in the pathogenesis of this bacterium [1]
The pathogenesis of bacterium is associated with its ability to invade macrophages and to circumvent bactericidal functions of the host cell in order to survive within a protective niche
We investigated the cellular and molecular mechanisms of DIM and demonstrated that DIM participate in the receptordependent phagocytosis of Mtb in human macrophages through a mechanism involving a reorganization of the plasma membrane following recognition of bacilli

Summary

Introduction

The cell envelope of Mycobacterium tuberculosis (Mtb) plays a major role in the pathogenesis of this bacterium [1]. DIM are produced by all members of the Mtb complex and a few other mycobacterial species, most pathogenic in humans or animals They are produced by the combined action of fatty acid synthases and polyketide synthases and in Mtb are composed of a mixture of long-chain b-diols esterified by multimethyl-branched fatty acids named mycocerosic acids [3]. An avirulent strain of Mtb coated with a mixture of DIM and cholesteryl oleate persisted longer than the uncoated strain in the spleen and lung of infected mice [8] Consistent with these initial observations, two independent signature-tagged transposon mutagenesis studies have led to the isolation of Mtb mutants with a severe growth defect in mice [9,10]. It remains unclear whether DIM mediate pathogenesis indirectly by changing the cell wall envelope or whether they act as effectors modifying host immune responses and notably interfering with antimicrobial activity

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Conclusion