Abstract
PE_PGRS proteins are unique to the Mycobacterium tuberculosis complex and a number of other pathogenic mycobacteria. PE_PGRS30, which is required for the full virulence of M. tuberculosis (Mtb), has three main domains, i.e. an N-terminal PE domain, repetitive PGRS domain and the unique C-terminal domain. To investigate the role of these domains, we expressed a GFP-tagged PE_PGRS30 protein and a series of its functional deletion mutants in different mycobacterial species (Mtb, Mycobacterium bovis BCG and Mycobacterium smegmatis) and analysed protein localization by confocal microscopy. We show that PE_PGRS30 localizes at the mycobacterial cell poles in Mtb and M. bovis BCG but not in M. smegmatis and that the PGRS domain of the protein strongly contributes to protein cellular localization in Mtb. Immunofluorescence studies further showed that the unique C-terminal domain of PE_PGRS30 is not available on the surface, except when the PGRS domain is missing. Immunoblot demonstrated that the PGRS domain is required to maintain the protein strongly associated with the non-soluble cellular fraction. These results suggest that the repetitive GGA-GGN repeats of the PGRS domain contain specific sequences that contribute to protein cellular localization and that polar localization might be a key step in the PE_PGRS30-dependent virulence mechanism.
Highlights
Despite many research and sanitary efforts, tuberculosis (TB) remains one of the deadliest human infectious diseases far from being defeated [1]
Polarization of PE_PGRS30GFP was found even more pronounced under this condition, while MtbPE_PGRS33GFP showed a diffused and homogenous fluorescence throughout the cell. Since their identification in the Mycobacterium tuberculosis (Mtb) genome, PE_PGRS proteins have been implicated in the mechanism of pathogenesis of TB and included in an hypothetical panel of surface mycobacterial antigens involved in immune evasion strategies [15,16,17]
In this study, using a panel of green fluorescent protein (GFP)-tagged proteins, we investigate the localization of PE_PGRS30 in three mycobacterial species (Mtb, M. bovis BCG and M. smegmatis) and analyzed the impact of the different protein domains on protein polarization on the bacterial cells
Summary
Despite many research and sanitary efforts, tuberculosis (TB) remains one of the deadliest human infectious diseases far from being defeated [1]. The poor knowledge of the biology of its causative agent, Mycobacterium tuberculosis (Mtb), is a main obstacle toward the development of improved control strategies [2,3] In this context, a better understanding of surface exposed, secreted and cell wall associated proteins is classically a key step to dissect the mechanisms of pathogenesis of bacteria and to identify antigens that may serve as candidate vaccines [4,5]. The recent discovery of the ESX secretion systems is shedding light on the mechanism whereby Mtb translocate effector proteins that are secreted or exposed on its surface and that can interfere with host components [9,10,11,12] The results of these studies are leading to the development of new vaccines and drug targets [13,14], emphasizing the impact that this line of research may have in the control of TB
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