Neha Quadir

Bio: Neha Quadir is an academic researcher from Indian Council of Medical Research. The author has contributed to research in topics: Autophagy & Innate immune system. The author has an hindex of 2, co-authored 4 publications receiving 9 citations. Previous affiliations of Neha Quadir include Jamia Hamdard.

Mycobacterium tuberculosis RipA Dampens TLR4-Mediated Host Protective Response Using a Multi-Pronged Approach Involving Autophagy, Apoptosis, Metabolic Repurposing, and Immune Modulation.

Abstract: Reductive evolution has endowed Mycobacterium tuberculosis (M. tb) with moonlighting in protein functions. We demonstrate that RipA (Rv1477), a peptidoglycan hydrolase, activates the NFκB signaling pathway and elicits the production of pro-inflammatory cytokines, TNF-α, IL-6, and IL-12, through the activation of an innate immune-receptor, toll-like receptor (TLR)4. RipA also induces an enhanced expression of macrophage activation markers MHC-II, CD80, and CD86, suggestive of M1 polarization. RipA harbors LC3 (Microtubule-associated protein 1A/1B-light chain 3) motifs known to be involved in autophagy regulation and indeed alters the levels of autophagy markers LC3BII and P62/SQSTM1 (Sequestosome-1), along with an increase in the ratio of P62/Beclin1, a hallmark of autophagy inhibition. The use of pharmacological agents, rapamycin and bafilomycin A1, reveals that RipA activates PI3K-AKT-mTORC1 signaling cascade that ultimately culminates in the inhibition of autophagy initiating kinase ULK1 (Unc-51 like autophagy activating kinase). This inhibition of autophagy translates into efficient intracellular survival, within macrophages, of recombinant Mycobacterium smegmatis expressing M. tb RipA. RipA, which also localizes into mitochondria, inhibits the production of oxidative phosphorylation enzymes to promote a Warburg-like phenotype in macrophages that favors bacterial replication. Furthermore, RipA also inhibited caspase-dependent programed cell death in macrophages, thus hindering an efficient innate antibacterial response. Collectively, our results highlight the role of an endopeptidase to create a permissive replication niche in host cells by inducing the repression of autophagy and apoptosis, along with metabolic reprogramming, and pointing to the role of RipA in disease pathogenesis.

Mycobacterium tuberculosis Protein PE6 (Rv0335c), a Novel TLR4 Agonist, Evokes an Inflammatory Response and Modulates the Cell Death Pathways in Macrophages to Enhance Intracellular Survival.

Abstract: Mycobacterium tuberculosis (M. tb) is an intracellular pathogen that exploits moonlighting functions of its proteins to interfere with host cell functions. PE/PPE proteins utilize host inflammatory signaling and cell death pathways to promote pathogenesis. We report that M. tb PE6 protein (Rv0335c) is a secretory protein effector that interacts with innate immune toll-like receptor TLR4 on the macrophage cell surface and promotes activation of the canonical NFĸB signaling pathway to stimulate secretion of proinflammatory cytokines TNF-α, IL-12, and IL-6. Using mouse macrophage TLRs knockout cell lines, we demonstrate that PE6 induced secretion of proinflammatory cytokines dependent on TLR4 and adaptor Myd88. PE6 possesses nuclear and mitochondrial targeting sequences and displayed time-dependent differential localization into nucleus/nucleolus and mitochondria, and exhibited strong Nucleolin activation. PE6 strongly induces apoptosis via increased production of pro-apoptotic molecules Bax, Cytochrome C, and pcMyc. Mechanistic details revealed that PE6 activates Caspases 3 and 9 and induces endoplasmic reticulum-associated unfolded protein response pathways to induce apoptosis through increased production of ATF6, Chop, BIP, eIF2α, IRE1α, and Calnexin. Despite being a potent inducer of apoptosis, PE6 suppresses innate immune defense strategy autophagy by inducing inhibitory phosphorylation of autophagy initiating kinase ULK1. Inversely, PE6 induces activatory phosphorylation of autophagy master regulator MtorC1, which is reflected by lower conversion of autophagy markers LC3BI to LC3BII and increased accumulation of autophagy substrate p62 which is also dependent on innate immune receptor TLR4. The use of pharmacological agents, rapamycin and bafilomycin A1, confirms the inhibitory effect of PE6 on autophagy, evidenced by the reduced conversion of LC3BI to LC3BII and increased accumulation of p62 in the presence of rapamycin and bafilomycin A1. We also observed that PE6 binds DNA, which could have significant implications in virulence. Furthermore, our analyses reveal that PE6 efficiently binds iron to likely aid in intracellular survival. Recombinant Mycobacterium smegmatis (M. smegmatis) containing pe6 displayed robust growth in iron chelated media compared to vector alone transformed cells, which suggests a role of PE6 in iron acquisition. These findings unravel novel mechanisms exploited by PE6 protein to subdue host immunity, thereby providing insights relevant to a better understanding of host-pathogen interaction during M. tb infection.

Post translational modifications in tuberculosis: ubiquitination paradox

Abstract: Innate immune signaling and xenophagy are crucial innate defense strategies exploited by the host to counteract intracellular pathogens with ubiquitination as a critical regulator of these processes. These pathogens, including Mycobacterium tuberculosis (M. tb), co-opt the host ubiquitin machinery by utilizing secreted or cell surface effectors to dampen innate host defenses. Inversely, the host utilizes ubiquitin ligase-mediated ubiquitination of intracellular pathogens and recruits autophagy receptors to induce xenophagy. In the current article, we discuss the co-option of the ubiquitin pathway by the M. tb virulence effectors.Abbreviations: ANAPC2: anaphase promoting complex subunit 2; IL: interleukin; Lys: lysine (K); MAPK: mitogen-activated protein kinase; MAP3K7/TAK1; mitogen-activated protein kinase kinase kinase 7; M. tb: Mycobacterium tuberculosis; NFKB/NF-κB: nuclear factor kappa B subunit; PtpA: protein tyrosine phosphatase; SQSTM1/p62: sequestosome 1; V-ATPase: vacuolar-type H+-ATPase; UBA: a eukaryotic-like ubiquitin-associated domain.

Mycobacterium tuberculosis RipA Dampens TLR4-Mediated Host Protective Response Using a Multi-Pronged Approach Involving Autophagy, Apoptosis, Metabolic Repurposing, and Immune Modulation.

Abstract: Reductive evolution has endowed Mycobacterium tuberculosis (M. tb) with moonlighting in protein functions. We demonstrate that RipA (Rv1477), a peptidoglycan hydrolase, activates the NFκB signaling pathway and elicits the production of pro-inflammatory cytokines, TNF-α, IL-6, and IL-12, through the activation of an innate immune-receptor, toll-like receptor (TLR)4. RipA also induces an enhanced expression of macrophage activation markers MHC-II, CD80, and CD86, suggestive of M1 polarization. RipA harbors LC3 (Microtubule-associated protein 1A/1B-light chain 3) motifs known to be involved in autophagy regulation and indeed alters the levels of autophagy markers LC3BII and P62/SQSTM1 (Sequestosome-1), along with an increase in the ratio of P62/Beclin1, a hallmark of autophagy inhibition. The use of pharmacological agents, rapamycin and bafilomycin A1, reveals that RipA activates PI3K-AKT-mTORC1 signaling cascade that ultimately culminates in the inhibition of autophagy initiating kinase ULK1 (Unc-51 like autophagy activating kinase). This inhibition of autophagy translates into efficient intracellular survival, within macrophages, of recombinant Mycobacterium smegmatis expressing M. tb RipA. RipA, which also localizes into mitochondria, inhibits the production of oxidative phosphorylation enzymes to promote a Warburg-like phenotype in macrophages that favors bacterial replication. Furthermore, RipA also inhibited caspase-dependent programed cell death in macrophages, thus hindering an efficient innate antibacterial response. Collectively, our results highlight the role of an endopeptidase to create a permissive replication niche in host cells by inducing the repression of autophagy and apoptosis, along with metabolic reprogramming, and pointing to the role of RipA in disease pathogenesis.

Mycobacterium tuberculosis Protein PE6 (Rv0335c), a Novel TLR4 Agonist, Evokes an Inflammatory Response and Modulates the Cell Death Pathways in Macrophages to Enhance Intracellular Survival.

Abstract: Mycobacterium tuberculosis (M. tb) is an intracellular pathogen that exploits moonlighting functions of its proteins to interfere with host cell functions. PE/PPE proteins utilize host inflammatory signaling and cell death pathways to promote pathogenesis. We report that M. tb PE6 protein (Rv0335c) is a secretory protein effector that interacts with innate immune toll-like receptor TLR4 on the macrophage cell surface and promotes activation of the canonical NFĸB signaling pathway to stimulate secretion of proinflammatory cytokines TNF-α, IL-12, and IL-6. Using mouse macrophage TLRs knockout cell lines, we demonstrate that PE6 induced secretion of proinflammatory cytokines dependent on TLR4 and adaptor Myd88. PE6 possesses nuclear and mitochondrial targeting sequences and displayed time-dependent differential localization into nucleus/nucleolus and mitochondria, and exhibited strong Nucleolin activation. PE6 strongly induces apoptosis via increased production of pro-apoptotic molecules Bax, Cytochrome C, and pcMyc. Mechanistic details revealed that PE6 activates Caspases 3 and 9 and induces endoplasmic reticulum-associated unfolded protein response pathways to induce apoptosis through increased production of ATF6, Chop, BIP, eIF2α, IRE1α, and Calnexin. Despite being a potent inducer of apoptosis, PE6 suppresses innate immune defense strategy autophagy by inducing inhibitory phosphorylation of autophagy initiating kinase ULK1. Inversely, PE6 induces activatory phosphorylation of autophagy master regulator MtorC1, which is reflected by lower conversion of autophagy markers LC3BI to LC3BII and increased accumulation of autophagy substrate p62 which is also dependent on innate immune receptor TLR4. The use of pharmacological agents, rapamycin and bafilomycin A1, confirms the inhibitory effect of PE6 on autophagy, evidenced by the reduced conversion of LC3BI to LC3BII and increased accumulation of p62 in the presence of rapamycin and bafilomycin A1. We also observed that PE6 binds DNA, which could have significant implications in virulence. Furthermore, our analyses reveal that PE6 efficiently binds iron to likely aid in intracellular survival. Recombinant Mycobacterium smegmatis (M. smegmatis) containing pe6 displayed robust growth in iron chelated media compared to vector alone transformed cells, which suggests a role of PE6 in iron acquisition. These findings unravel novel mechanisms exploited by PE6 protein to subdue host immunity, thereby providing insights relevant to a better understanding of host-pathogen interaction during M. tb infection.

The Role of TLR2 in Infectious Diseases Caused by Mycobacteria: From Cell Biology to Therapeutic Target

Abstract: Simple Summary Due to the broad functions of Toll-like receptor 2 (TLR2) in innate immunity, the drive for the development of TLR2-targeted therapeutic treatments has accelerated in recent decades. However, its dual role in both the activation and suppression of innate immune responses makes it very difficult to use the results from gathered basic research and apply them to the development of clinical trials. Therefore, this review aims to summarize the knowledge of the function of TLR2 in innate immunity and metabolism to provide some future research directions. Abstract Innate immunity is considered the first line of defense against microbial invasion, and its dysregulation can increase the susceptibility of hosts to infections by invading pathogens. Host cells rely on pattern recognition receptors (PRRs) to recognize invading pathogens and initiate protective innate immune responses. Toll-like receptor 2 (TLR2) is believed to be among the most important Toll-like receptors for defense against mycobacterial infection. TLR2 has been reported to have very broad functions in infectious diseases and also in other diseases, such as chronic and acute inflammatory diseases, cancers, and even metabolic disorders. However, TLR2 has an unclear dual role in both the activation and suppression of innate immune responses. Moreover, in some studies, the function of TLR2 was shown to be controversial, and therefore its role in several diseases is still inconclusive. Therefore, although TLR2 has been shown to have an important function in innate immunity, its usefulness as a therapeutic target in clinical application is still uncertain. In this literature review, we summarize the knowledge of the functions of TLR2 in host–mycobacterial interactions, discuss controversial results, and suggest possibilities for future research.

The exploitation of host autophagy and ubiquitin machinery by Mycobacterium tuberculosis in shaping immune responses and host defense during infection

Abstract: ABSTRACT Intracellular pathogens have evolved various efficient molecular armaments to subvert innate defenses. Cellular ubiquitination, a normal physiological process to maintain homeostasis, is emerging one such exploited mechanism. Ubiquitin (Ub), a small protein modifier, is conjugated to diverse protein substrates to regulate many functions. Structurally diverse linkages of poly-Ub to target proteins allow enormous functional diversity with specificity being governed by evolutionarily conserved enzymes (E3-Ub ligases). The Ub-binding domain (UBD) and LC3-interacting region (LIR) are critical features of macroautophagy/autophagy receptors that recognize Ub-conjugated on protein substrates. Emerging evidence suggests that E3-Ub ligases unexpectedly protect against intracellular pathogens by tagging poly-Ub on their surfaces and targeting them to phagophores. Two E3-Ub ligases, PRKN and SMURF1, provide immunity against Mycobacterium tuberculosis (M. tb). Both enzymes conjugate K63 and K48-linked poly-Ub to M. tb for successful delivery to phagophores. Intriguingly, M. tb exploits virulence factors to effectively dampen host-directed autophagy utilizing diverse mechanisms. Autophagy receptors contain LIR-motifs that interact with conserved Atg8-family proteins to modulate phagophore biogenesis and fusion to the lysosome. Intracellular pathogens have evolved a vast repertoire of virulence effectors to subdue host-immunity via hijacking the host ubiquitination process. This review highlights the xenophagy-mediated clearance of M. tb involving host E3-Ub ligases and counter-strategy of autophagy inhibition by M. tb using virulence factors. The role of Ub-binding receptors and their mode of autophagy regulation is also explained. We also discuss the co-opting and utilization of the host Ub system by M. tb for its survival and virulence. Abbreviations: APC: anaphase promoting complex/cyclosome; ATG5: autophagy related 5; BCG: bacille Calmette-Guerin; C2: Ca2+-binding motif; CALCOCO2: calcium binding and coiled-coil domain 2; CUE: coupling of ubiquitin conjugation to ER degradation domains; DUB: deubiquitinating enzyme; GABARAP: GABA type A receptor-associated protein; HECT: homologous to the E6-AP carboxyl terminus; IBR: in-between-ring fingers; IFN: interferon; IL1B: interleukin 1 beta; KEAP1: kelch like ECH associated protein 1; LAMP1: lysosomal associated membrane protein 1; LGALS: galectin; LIR: LC3-interacting region; MAPK11/p38: mitogen-activated protein kinase 11; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MAP3K7/TAK1: mitogen-activated protein kinase kinase kinase 7; MAPK8/JNK: mitogen-activated protein kinase 8; MHC-II: major histocompatibility complex-II; MTOR: mechanistic target of rapamycin kinase; NBR1: NBR1 autophagy cargo receptor; NFKB1/p50: nuclear factor kappa B subunit 1; OPTN: optineurin; PB1: phox and bem 1; PE/PPE: proline-glutamic acid/proline-proline-glutamic acid; PknG: serine/threonine-protein kinase PknG; PRKN: parkin RBR E3 ubiquitin protein ligase; RBR: RING-in between RING; RING: really interesting new gene; RNF166: RING finger protein 166; ROS: reactive oxygen species; SMURF1: SMAD specific E3 ubiquitin protein ligase 1; SQSTM1: sequestosome 1; STING1: stimulator of interferon response cGAMP interactor 1; TAX1BP1: Tax1 binding protein 1; TBK1: TANK binding kinase 1; TNF: tumor necrosis factor; TRAF6: TNF receptor associated factor 6; Ub: ubiquitin; UBA: ubiquitin-associated; UBAN: ubiquitin-binding domain in ABIN proteins and NEMO; UBD: ubiquitin-binding domain; UBL: ubiquitin-like; ULK1: unc-51 like autophagy activating kinase 1.

Macrophage: A Cell With Many Faces and Functions in Tuberculosis

Abstract: Mycobacterium tuberculosis (Mtb) is the causative agent of human tuberculosis (TB) which primarily infects the macrophages. Nearly a quarter of the world’s population is infected latently by Mtb. Only around 5%–10% of those infected develop active TB disease, particularly during suppressed host immune conditions or comorbidity such as HIV, hinting toward the heterogeneity of Mtb infection. The aerosolized Mtb first reaches the lungs, and the resident alveolar macrophages (AMs) are among the first cells to encounter the Mtb infection. Evidence suggests that early clearance of Mtb infection is associated with robust innate immune responses in resident macrophages. In addition to lung-resident macrophage subsets, the recruited monocytes and monocyte-derived macrophages (MDMs) have been suggested to have a protective role during Mtb infection. Mtb, by virtue of its unique cell surface lipids and secreted protein effectors, can evade killing by the innate immune cells and preferentially establish a niche within the AMs. Continuous efforts to delineate the determinants of host defense mechanisms have brought to the center stage the crucial role of macrophage phenotypical variations for functional adaptations in TB. The morphological and functional heterogeneity and plasticity of the macrophages aid in confining the dissemination of Mtb. However, during a suppressed or hyperactivated immune state, the Mtb virulence factors can affect macrophage homeostasis which may skew to favor pathogen growth, causing active TB. This mini-review is aimed at summarizing the interplay of Mtb pathomechanisms in the macrophages and the implications of macrophage heterogeneity and plasticity during Mtb infection.