Mycobacterium Tuberculosis Region Of Difference (RD)1Protein PE35Perturbs IL-1βSignaling In A TLR2Dependent Manner

Mycobacterium Tuberculosis (Mtb), the etiological agent of tuberculosis, is one of the most successful pathogen, with approximately one-third of the world’s population infected, almost1.5million people killed annually around the world. The emergence of antibiotic-resistant strains of M. Tuberculosis and/or co-infection with HIV aggravates the risk for mortality.The PE gene family are only present in the genus Mycobacterium. The PE family genes are classified into two subfamilies:PE_and PE_PGRS. So far, the function of PE_PGRS protein is well studied, including altering the cellular structure and the colony morphology, serving as lipase to supply energy for persistent mycobacteria, involving in antigenic variation, and maintaining M. Tuberculosis persistence within granulomas. However, the pathophysiological functions of proline-glutamic acid (PE) family of proteins of M. Tuberculosis remained an elusive issue. PE35, a member of PE family, belonging to RD1, is conserved in virulent mycobacterium, absent in attenuated BCG, this may be related with virulence of MTB.The mycobacterial components were recognized by different pattern recognition receptors (PPRs) lining on innate immune cells, leading to the induction of cytokines, contributing to the early control of infection. Many cytokines participated in MTB control, such as IL-12p70, IL-23, IL-27, IL-35, IL-la, IL-1β, IL-18, IL-33,TNF-a, IFN-γ, Type I IFNs, IL-17, Th17-related cytokines, IL-10and other immunosuppressive cytokines.In this study, we mainly foucus on the role of PE35in proinflammatory cytokine production in macrophages, identifies the potential siganling pathway. Therefore, we amplified PE35from M. Tuberculosis H37Rv genome, the PCR products were ligated to the pMD19-T Simple Vector, and then subcloned into vector pEGFP-C1. The recombinant plasmid pEGFP-C1-PE35was transfected into RAW264.7cell. Pretreatment transfected RAW264.7cell with LPS, the cytokine production was assayed by real time PCR and ELISA. In addition, the phosphorylation level of p38MAPK and Erk1/2was assayed to identify the signaling cascade that cause decrease in cytokine production. Additionally, the interation of PE35with TLR2was carried by pull down.Here, we demonstrated that PE35attenuates LPS induced IL-1β production in transfected RAW264.7cell, which is independent of p38mitogen-activated protein kinase (MAPK) and extracellular signal-related kinases(ERK)1/2. Furthermore, pull down assay reveals the interaction of PE35with TLR2. IL-1β was showed to trigger a Th17bias in the cellular adaptive responses. These results collectively indicate that PE35may act as a novel TLR2ligand, attenuates the induction of Th17adaptive response by decreasing IL-1β release, contributing to the immune invasion of MTB.