Evaluation of PE_PGRS33 as a potential surface target for humoral responses against Mycobacterium tuberculosis

Abstract

Mycobacterium tuberculosis (Mtb) PE_PGRS33 is a surface-exposed protein that was shown to interact with Toll-like receptor 2 on host macrophages to induce inflammatory signals and promote entry in macrophages. In this study, we investigated PE_PGRS33 as a potential target of a humoral response aimed at hampering key processes in tuberculosis pathogenesis. PE_PGRS33 protein was successfully expressed and purified under native condition in Escherichia coli. The purified protein retained its native functional and biological properties, showing the ability to elicit proinflammatory signals in murine and human macrophages. Interestingly, a polyclonal antiserum raised against native PE_PGRS33 showed no cross-reactions with other mycobacterial proteins. The anti-PE_PGRS33 serum was also able to inhibit Mtb entry into macrophages, but it did not reduce entry of the MtbΔpe_pgrs33 strain. Addition of native recombinant PE_PGRS33 to the MtbΔpe_pgrs33 strain during infection restored the Mtb wild-type entry phenotype in macrophage. Moreover, the anti-PE_PGRS33 serum was able to neutralize the proinflammatory activity of PE_PGRS33 in vitro. Furthermore, mice immunized with native recombinant PE_PGRS33, but not with a DNA vaccine expressing PE_PGRS33, were able to restrict M. smegmatis in vivo. These results highlight the potential use of PE_PGRS33 as a target of a neutralizing humoral response against tuberculosis.

PE_PGRS proteins ofMycobacterium tuberculosis: A specialized molecular task force at the forefront of host–pathogen interaction

“…Of note, PE_PGRS33 was able to interact with TLR2 to promote cell death and inflammation [ 42 ] and proper localization of PE_PGRS33 on the mycobacterial surface is required to activate the TLR2 pathway [ 55 ]. Moreover, an antiserum directed against the native form of PE_PGRS33 was able to abolish the secretion of TNF-α following infection of macrophages with M. smegmatis expressing PE_PGRS33, further supporting the key role of PE_PGRS33-TLR2 interaction [ 81 ]. In line with these findings, the Mtb Δ pe_pgrs 33 mutant was impaired, compared to the parental strain, in its ability to enter in macrophages, but not epithelial cells, in a process involving activation of the TLR2-CR3 pathway, which activates the inside-out-signaling to promote Mtb entry in macrophages [ 44 ].…”

“…Heterologous expression of PE_PGRS33 in M. smegmatis promoted cell death and increased mycobacterial survival in macrophages and in intraperitoneally infected mice over the parental strain or the M. smegmatis recombinant strain expressing only the PE domain of PE_PGRS33, pointing for the key role of the PGRS domain in this process [ 42 , 78 , 79 ]. PE_PGRS33 triggered TNF-α and IL-12 secretion promoting cell necrosis and inflammation, as highlighted by the enlarged spleens of mice infected with M. smegmatis expressing PE_PGRS33 compared to controls [ 55 , 78 , 80 , 81 ]. However, experiments carried out with the purified recombinant protein or in eukaryotic cells transfected with a plasmid expressing PE_PGRS33, while confirming the ability of PE_PGRS33 to promote cell death implicated a mechanism involving apoptosis rather than necrosis [ 42 , 79 ].…”

Graphene oxide prevents mycobacteria entry into macrophages through extracellular entrapment

“…Conversely, Mtb infection was carried out using an MOI of 1 (1 : 1 bacterium to cell ratio) and the initial infection was for 1 hour instead of 4 hours, in line with our previous research. 50,51 In the third experimental setting, macrophages were pre-treated with medium alone and medium containing GO (1000 mg ml À1 , 100 mg ml À1 and 10 mg ml À1 ) or gentamycin (10 mg ml À1 ). Four hours later, the macrophages were infected with Ms GFP (MOI 10) or Mtb GFP (MOI 1) and CFUs were obtained as previously described.…”

PE_PGRS31-S100A9 Interaction Promotes Mycobacterial Survival in Macrophages Through the Regulation of NF-κB-TNF-α Signaling and Arachidonic Acid Metabolism

“…Our present results show that Rv1768 is located in the bacterial CW and only exits in H37Rv and H37Ra, not in M. avium, Ms, M. marinum, M. intracellulare, or BCG. Some other members of the M. tb PE_PGRS family, for example, PE_PGRS33, are surface-exposed proteins that interact with TLR2 on host macrophages to induce inflammatory signals and promote entry into macrophages (Minerva et al, 2017). PE_PGRS3 mediates adhesion and persistence in host cells (De Maio et al, 2018).…”