The tubercle bacillus parasitizes macrophages by inhibiting phagosome maturation into the phagolysosome. This phenomenon underlies the tuberculosis pandemic involving 2 billion people. We report here how Mycobacterium tuberculosis causes phagosome maturation arrest. A glycosylated M. tuberculosis phosphatidylinositol [mannose-capped lipoarabinomannan (ManLAM)] interfered with the phagosomal acquisition of the lysosomal cargo and syntaxin 6 from the trans-Golgi network. ManLAM specifically inhibited the pathway dependent on phosphatidylinositol 3-kinase activity and phosphatidylinositol 3-phosphate-binding effectors. These findings identify ManLAM as the M. tuberculosis product responsible for the inhibition of phagosomal maturation.

Live Mycobacterium tuberculosis persists in macrophage phagosomes by interfering with phagolysosome biogenesis. Here, using four-dimensional microscopy and in vitro assays, we report the principal difference between phagosomes containing live and dead mycobacteria. Phosphatidylinositol 3-phosphate (PI3P), a membrane trafficking regulatory lipid essential for phagosomal acquisition of lysosomal constituents, is retained on phagosomes harboring dead mycobacteria but is continuously eliminated from phagosomes with live bacilli. We show that the exclusion of PI3P from live mycobacterial phagosomes can be only transiently reversed by Ca 2؉ fluxes, and that live M. tuberculosis secretes a lipid phosphatase, SapM, that hydrolyzes PI3P, inhibits phagosome-late endosome fusion in vitro, and contributes to inhibition of phagosomal maturation.macrophage ͉ phagosome ͉ tuberculosis ͉ lysosome ͉ phosphatidylinositol 3-phosphate T he infectious cycle of Mycobacterium tuberculosis rests upon the ability of this potent pathogen to parasitize host mononuclear phagocytic cells (1). In infected macrophages, M. tuberculosis resides within a phagosome that avoids the default maturation pathway leading to phagolysosome formation (2). The salient characteristics of the mycobacterial phagosome include (i) paucity of vacuolar H ϩ ATPase (3), (ii) attendant inefficient luminal acidification (3); and (iii) inadequate levels of mature lysosomal hydrolases (3, 4). These and additional (4-6) properties of the M. tuberculosis phagosome promote intracellular survival and growth of the tubercle bacilli and help avoid their immunological detection (1).The arrest of M. tuberculosis phagosome maturation has been studied at the membrane-trafficking level (2), with a focus on the small GTP-binding proteins, including Rab GTPases (7-9). Rabs direct intracellular trafficking by regulating activity and recruitment to organellar membranes of Rab-interacting partners and downstream effectors (10). The initial analyses of Rabs on mycobacterial phagosomes have indicated that the M. tuberculosis phagolysosome biogenesis arrest occurs between the stages controlled by the early endosomal GTPase Rab5 and its late endosomal counterpart Rab7 (7). A number of follow-up studies have indicated critical contributions of Rab5 effectors in mycobacterial phagosome maturation arrest, with a prominent role for the phosphatidylinositol 3-kinase (PI3K) hVPS34, its product phosphatidylinositol 3-phosphate (PI3P), and an array of PI3P-binding proteins (11)(12)(13)(14). PI3P affects localization and function of proteins containing the PI3P-binding domains (FYVE, PH, and PX) (15). These proteins in turn execute various steps in membrane trafficking, endosomal protein sorting, and multisubunit enzyme assembly at the membrane, including phagosomal maturation (11, 16), early endosomal homotypic fusion (17), delivery of internalized plasma membrane receptors to late endosomes (18), formation of internal vesicles within late endosomal multivesicular bodies involved in termination of...

Phagocytosis and phagolysosome biogenesis represent fundamental biological processes essential for proper tissue homeostasis, development, elimination of invading microorganisms, and antigen processing and presentation. Phagosome formation triggers a preprogrammed pathway of maturation into the phagolysosome, a process controlled by Ca2+ and the regulators of organellar trafficking centered around the small GTP-binding proteins Rabs and their downstream effectors, including lipid kinases, organellar tethering molecules, and membrane fusion apparatus. Mycobacterium tuberculosis is a potent human pathogen parasitizing macrophages. It interferes with the Rab-controlled membrane trafficking and arrests the maturing phagosome at a stage where no harm can be done to the pathogen while the delivery of nutrients and membrane to the vacuole harboring the microorganism continues. This process, referred to as the M. tuberculosis phagosome maturation arrest or inhibition of phagosome-lysosome fusion, is critical for M. tuberculosis persistence in human populations. It also provides a general model system for dissecting the phagolysosome biogenesis pathways. Here we review the fundamental trafficking processes targeted by M. tuberculosis and the mycobacterial products that interfere with phagosomal maturation.