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The misfolding and mutation of Cu/Zn superoxide dismutase (SOD1) is commonly associated with amyotrophic lateral sclerosis (ALS). SOD1 can accumulate within stress granules (SGs), a type of membraneless organelle, which is believed to form via liquid–liquid phase separation (LLPS). Using wild‐type, metal‐deficient, and different ALS disease mutants of SOD1 and computer simulations, we report here that the absence of Zn leads to structural disorder within two loop regions of SOD1, triggering SOD1 LLPS and amyloid formation. The addition of exogenous Zn to either metal‐free SOD1 or to the severe ALS mutation I113T leads to the stabilization of the loops and impairs SOD1 LLPS and aggregation. Moreover, partial Zn‐mediated inhibition of LLPS was observed for another severe ALS mutant, G85R, which shows perturbed Zn‐binding. By contrast, the ALS mutant G37R, which shows reduced Cu‐binding, does not undergo LLPS. In addition, SOD1 condensates induced by Zn‐depletion exhibit greater cellular toxicity than aggregates formed by prolonged incubation under aggregating conditions. Overall, our work establishes a role for Zn‐dependent modulation of SOD1 conformation and LLPS properties that may contribute to amyloid formation. Synopsis: The formation of membraneless organelles, such as stress granules (SGs), is believed to occur through the process of liquid–liquid phase separation (LLPS). The amyotrophic lateral sclerosis (ALS) disease‐associated protein SOD1 is known to accumulate within SGs, but if it itself undergoes LLPS, and if this affects amyloid formation, is not well‐studied. The apo form of SOD1 undergoes LLPS.SOD1 LLPS and subsequent aggregation are regulated by a conformational transition between a disordered and a relatively compact folded state.The metal cofactor Zn prevents SOD1 LLPS via a conformational transition.SOD1 condensates show greater cytotoxicity than aggregates. [ABSTRACT FROM AUTHOR]

HuR is a miRNA derepressor protein that can act as miRNA sponge for specific miRNAs to negate their action on target mRNAs. Here we have identified how HuR, by inducing extracellular vesicles‐mediated export of miRNAs, ensures robust derepression of miRNA‐repressed cytokines essential for strong pro‐inflammatory response in activated mammalian macrophages. Leishmania donovani, the causative agent of visceral leishmaniasis, on the contrary alters immune response of the host macrophage by a variety of complex mechanisms to promote anti‐inflammatory response essential for the survival of the parasite. We have found that during Leishmania infection, the pathogen targets HuR to promote onset of anti‐inflammatory response in mammalian macrophages. In infected macrophages, Leishmania also upregulate protein phosphatase 2A that acts on Ago2 protein to keep it in dephosphorylated and miRNA‐associated form. This causes robust repression of the miRNA‐targeted pro‐inflammatory cytokines to establish an anti‐inflammatory response in infected macrophages. HuR has an inhibitory effect on protein phosphatase 2A expression, and mathematical modelling of macrophage activation process supports antagonistic miRNA‐modulatory roles of HuR and protein phosphatase 2A which mutually balances immune response in macrophage by targeting miRNA function. Supporting this model, ectopic expression of the protein HuR and simultaneous inhibition of protein phosphatase 2A induce strong pro‐inflammatory response in the host macrophage to prevent the virulent antimonial drug‐sensitive or drug‐resistant form of L. donovani infection. Thus, HuR can act as a balancing factor of immune responses to curtail the macrophage infection process by the protozoan parasite. Synopsis: Downregulation of HuR and upregulation of PP2A phosphatase are key mechanisms by which Leishmania donovani modulates the inflammatory response in host macrophage. Simultaneously inducing HuR and inhibiting PP2A could to be new way of treating the drug resistant form of the disease. PP2A promotes anti‐inflammatory response by promoting dephosphorylation and miRNA re‐association of Ago2.HuR induces pro‐inflammatory response in macrophage by favouring extracellular export of Ago2 uncoupled miRNAs.Leishmania donovani downregulates HuR and upregulates PP2A to establish anti‐ inflammatory response in infected cells.HuR clears macrophage internalized pathogens both in ex vivo and in vivo context. [ABSTRACT FROM AUTHOR]

MPT63, a major secreted protein from Mycobacterium tuberculosis, has been shown to have immunogenic properties and has been implicated in virulence. MPT63 is a β-sandwich protein containing 11 β strands and a very short stretch of 310 helix. The detailed experimental and computational study reported here investigates the equilibrium unfolding transition of MPT63. It is shown that in spite of being a complete β-sheet protein, MPT63 has a strong propensity toward helix structures in its early intermediates. Far UV-CD and FTIR spectra clearly suggest that the low-pH intermediate of MTP63 has enhanced helical content, while fluorescence correlation spectroscopy suggests a significant contraction. Molecular dynamics simulation complements the experimental results indicating that the unfolded state of MPT63 traverses through intermediate forms with increased helical characteristics. It is found that this early intermediate contains exposed hydrophobic surface, and is aggregation prone. Although MPT63 is a complete β-sheet protein in its native form, the present findings suggest that the secondary structure preferences of the local interactions in early folding pathway may not always follow the native conformation. Furthermore, the Gly25Ala mutant supports the proposed hypothesis by increasing the non-native helical propensity of the protein structure. [ABSTRACT FROM AUTHOR]