Proteomic characterization of the Mycobacterium marinum-containing vacuole in Dictyostelium discoideum

Abstract

Mycobacterium tuberculosis, the causative agent of tuberculosis, is able to manipulate the phagosome compartment in which it resides in order to establish a permissive replicative compartment called the Mycobacterium-containing vacuole (MCV). Mycobacterium marinum, a fish pathogen and a close relative of the tuberculosis group, is able to infect the free-living amoeba and professional phagocyte Dictyostelium discoideum and to manipulate its phagosome maturation. By using this host-pathogen model system, we have established an innovative process to isolate MCVs. This procedure allowed us to isolate the M. marinum-MCV at 1, 3 and 6 hours post infection to study the early M. marinum-MCV proteome. By using isobaric labelling and mass spectrometry, we quantitatively compared the proteomic composition of those MCVs isolated at different stages of the early infection phase to understand how M. marinum impacts on this compartment to divert it from the normal phagosomal pathway. Furthermore, we also compared the manipulated compartment M. marinum-MCV to non- or less manipulated compartments containing different mycobacteria strains: the non-pathogenic M. smegmatis, the avirulent M. marinum-L1D or the attenuated M. marinum-RD1.