Systematic Identification of Mycobacterium tuberculosis Effectors Reveals that BfrB Suppresses Innate Immunity

Abstract

(Mtb) has evolved multiple strategies to counter the human immune system. The effectors of Mtb play important roles in the interactions with the host. However, because of the lack of highly efficient strategies, there are only a handful of known Mtb effectors, thus hampering our understanding of Mtb pathogenesis. In this study, we probed Mtb proteome microarray with biotinylated whole-cell lysates of human macrophages, identifying 26 Mtb membrane proteins and secreted proteins that bind to macrophage proteins. Combining GST pull-down with mass spectroscopy then enabled the specific identification of all binders. We refer to this proteome microarray-based strategy as SOPHIE (Systematic unlOcking of Pathogen and Host Interacting Effectors). Detailed investigation of a novel effector identified here, the iron storage protein BfrB (Rv3841), revealed that BfrB inhibits NF-κB-dependent transcription through binding and reducing the nuclear abundance of the ribosomal protein S3 (RPS3), which is a functional subunit of NF- κB. The importance of this interaction was evidenced by the promotion of survival in macrophages of the mycobacteria, by overexpression of BfrB. Thus, beyond demonstrating the power of SOPHIE in the discovery of novel effectors of human pathogens, we expect that the set of Mtb effectors identified in this work will greatly facilitate the understanding of the pathogenesis of Mtb, possibly leading to additional potential molecular targets in the battle against tuberculosis.

The Nitrogen Regulator GlnR Directly Controls Transcription of the prpDBC Operon Involved in Methylcitrate Cycle in Mycobacterium smegmatis

“…The human mononuclear macrophage THP-1 cell was from the Cell Bank of Typical Culture Preservation Committee of Chinese Academy of Science (Shanghai, China) and was cultured in RPMI 1640 medium with 10% fetal bovine serum (FBS) at 37°C in the presence of 5% CO 2 . They were divided into 12-well plates (2.0 ϫ 10 5 cells per well) and differentiated using phorbol myristate acetate (PMA) when the cells were at the best state (41). After 12 h, the cells were washed and then cultured in fresh RPMI 1640 medium for 12 h. The M. smegmatis bacteria were added into the plates (10 times more than the THP-1 cells).…”

Nitrogen regulator GlnR directly controls transcription ofprpDBCoperon involved in methylcitrate cycle inMycobacterium smegmatis

“…The human mononuclear macrophage THP-1 cell was from the Cell Bank of Typical Culture Preservation Committee of Chinese Academy of Science (Shanghai, China), and was cultured in RPMI-1640 medium with 10% FBS at 37 °C in the presence of 5% CO 2 . They were divided into 12-well plates (2.0 × 10 5 cells per well) and differentiated using PMA when the cells were at the best state [23]. After 12 h, the cells were washed and then cultured in fresh RPMI-1640 medium for 12 h. The M. smegmatis were added into the plates (ten times more than the THP-1 cells).…”

A pathway‐based association analysis identified FMNL1‐MAP3K14 as susceptibility genes for leprosy

“…Leprosy is a disease whose clinical reaction depends on the cytokine produced during the immune response . The transcription factor NF‐κB has a critical role in coordinating the expression of a wide variety of genes that regulate innate immunity by controlling the expression of various immune regulatory molecules and thus modulates the intracellular survival of pathogens . Therefore, particular association study to confirm the relationship between leprosy and NF‐kB pathway is necessary…”

“…The colour of SNP is equivalent to the strength of linkage disequilibrium (r 2 ) to the top SNP variety of genes that regulate innate immunity by controlling the expression of various immune regulatory molecules and thus modulates the intracellular survival of pathogens. [26,27] Therefore, particular association study to confirm the relationship between leprosy and NF-kB pathway is necessary. [25] M. leprae mainly triggers the TLR2 signalling pathway [28] and subsequently activates macrophages that can increase the release of inflammatory factors, such as TNFα and IL-1, which will enhance both the nuclear translocation and the DNA-binding activity of NF-κB.…”