P38 and JNK Mitogen-Activated Protein Kinases Interact With Chikungunya Virus Non-structural Protein-2 and Regulate TNF Induction During Viral Infection in Macrophages

Chikungunya virus (CHIKV), a mosquito-borne Alphavirus, is endemic in different parts of the globe. The host macrophages are identified as the major cellular reservoirs of CHIKV during infection and this virus triggers robust TNF production in the host macrophages, which might be a key mediator of virus induced inflammation. However, the molecular mechanism underneath TNF induction is not understood yet. Accordingly, the Raw264.7 cells, a mouse macrophage cell line, were infected with CHIKV to address the above-mentioned question. It was observed that CHIKV induces both p38 and JNK phosphorylation in macrophages in a time-dependent manner and p-p38 inhibitor, SB203580 is effective in reducing infection even at lower concentration as compared to the p-JNK inhibitor, SP600125. However, inhibition of p-p38 and p-JNK decreased CHIKV induced TNF production in the host macrophages. Moreover, CHIKV induced macrophage derived TNF was found to facilitate TCR driven T cell activation. Additionally, it was noticed that the expressions of key transcription factors involved mainly in antiviral responses (p-IRF3) and TNF production (p-c-jun) were induced significantly in the CHIKV infected macrophages as compared to the corresponding mock cells. Further, it was demonstrated that CHIKV mediated TNF production in the macrophages is dependent on p38 and JNK MAPK pathways linking p-c-jun transcription factor. Interestingly, it was found that CHIKV nsP2 interacts with both p-p38 and p-JNK MAPKs in the macrophages. This observation was supported by the in silico protein-protein docking analysis which illustrates the specific amino acids responsible for the nsP2-MAPKs interactions. A strong polar interaction was predicted between Thr-180 (within the phosphorylation lip) of p38 and Gln-273 of nsP2, whereas, no such polar interaction was predicted for the phosphorylation lip of JNK which indicates the differential roles of p-p38 and p-JNK during CHIKV infection in the host macrophages. In summary, for the first time it has been shown that CHIKV triggers robust TNF production in the host macrophages via both p-p38 and p-JNK/p-c-jun pathways and the interaction of viral protein, nsP2 with these MAPKs during infection. Hence, this information might shed light in rationale-based drug designing strategies toward a possible control measure of CHIKV infection in future

Oxidation Induced Doping of Nanoparticles Revealed by in Situ X-ray Absorption Studies

Doping is a well-known approach to modulate the electronic and optical properties of nanoparticles (NPs). However, doping at nanoscale is still very challenging, and the reasons for that are not well understood. We studied the formation and doping process of iron and iron oxide NPs in real time by in situ synchrotron X-ray absorption spectroscopy. Our study revealed that the mass flow of the iron triggered by oxidation is responsible for the internalization of the dopant (molybdenum) adsorbed at the surface of the host iron NPs. The oxidation induced doping allows controlling the doping levels by varying the amount of dopant precursor. Our in situ studies also revealed that the dopant precursor substantially changes the reaction kinetics of formation of iron and iron oxide NPs. Thus, in the presence of dopant precursor we observed significantly faster decomposition rate of iron precursors and substantially higher stability of iron NPs against oxidation. The same doping mechanism and higher stability of host metal NPs against oxidation was observed for cobalt-based systems. Since the internalization of the adsorbed dopant at the surface of the host NPs is driven by the mass transport of the host, this mechanism can be potentially applied to introduce dopants into different oxidized forms of metal and metal alloy NPs providing the extra degree of compositional control in material design.Fil: Kwon, Soon Gu. Argonne National Laboratory; Estados UnidosFil: Chattopadhyay, Soma. Argonne National Laboratory; Estados Unidos. Illinois Institute of Technology; Estados UnidosFil: Koo, Bonil. Argonne National Laboratory; Estados UnidosFil: Dos Santos Claro, Paula Cecilia. Argonne National Laboratory; Estados UnidosFil: Shibata, Tomohiro. Argonne National Laboratory; Estados UnidosFil: Requejo, Felix Gregorio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Fisicoquímicas Teóricas y Aplicadas. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Investigaciones Fisicoquímicas Teóricas y Aplicadas; ArgentinaFil: Giovanetti, Lisandro Jose. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Fisicoquímicas Teóricas y Aplicadas. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Investigaciones Fisicoquímicas Teóricas y Aplicadas; ArgentinaFil: Liu, Yuzi. Argonne National Laboratory; Estados UnidosFil: Johnson, Christopher. Argonne National Laboratory; Estados UnidosFil: Prakapenka, Vitali. University of Chicago; Estados UnidosFil: Lee, Byeongdu. Argonne National Laboratory; Estados UnidosFil: Shevchenko, Elena V.. Argonne National Laboratory; Estados Unido

Inhibition of IRGM establishes a robust antiviral immune state to restrict pathogenic viruses

The type I interferon (IFN) response is the major host arsenal against invading viruses. IRGM is a negative regulator of IFN responses under basal conditions. However, the role of human IRGM during viral infection has remained unclear. In this study, we show that IRGM expression is increased upon viral infection. IFN responses induced by viral PAMPs are negatively regulated by IRGM. Conversely, IRGM depletion results in a robust induction of key viral restriction factors including IFITMs, APOBECs, SAMHD1, tetherin, viperin, and HERC5/6. Additionally, antiviral processes such as MHC-I antigen presentation and stress granule signaling are enhanced in IRGM-deficient cells, indicating a robust cell-intrinsic antiviral immune state. Consistently, IRGM-depleted cells are resistant to the infection with seven viruses from five different families, including Togaviridae, Herpesviridae, Flaviviverdae, Rhabdoviridae, and Coronaviridae. Moreover, we show that Irgm1 knockout mice are highly resistant to chikungunya virus (CHIKV) infection. Altogether, our work highlights IRGM as a broad therapeutic target to promote defense against a large number of human viruses, including SARS-CoV-2, CHIKV, and Zika virus

TLR4 is one of the receptors for Chikungunya virus envelope protein E2 and regulates virus induced pro-inflammatory responses in host macrophages

Toll like receptor 4 (TLR4), a pathogen-associated molecular pattern (PAMP) receptor, is known to exert inflammation in various cases of microbial infection, cancer and autoimmune disorders. However, any such involvement of TLR4 in Chikungunya virus (CHIKV) infection is yet to be explored. Accordingly, the role of TLR4 was investigated towards CHIKV infection and modulation of host immune responses in the current study using mice macrophage cell line RAW264.7, primary macrophage cells of different origins and in vivo mice model. The findings suggest that TLR4 inhibition using TAK-242 (a specific pharmacological inhibitor) reduces viral copy number as well as reduces the CHIKV-E2 protein level significantly using p38 and JNK-MAPK pathways. Moreover, this led to reduced expression of macrophage activation markers like CD14, CD86, MHC-II and pro-inflammatory cytokines (TNF, IL-6, MCP-1) significantly in both the mouse primary macrophages and RAW264.7 cell line, in vitro. Additionally, TAK-242-directed TLR4 inhibition demonstrated a significant reduction of percent E2-positive cells, viral titre and TNF expression in hPBMC-derived macrophages, in vitro. These observations were further validated in TLR4-knockout (KO) RAW cells. Furthermore, the interaction between CHIKV-E2 and TLR4 was demonstrated by immuno-precipitation studies, in vitro and supported by molecular docking analysis, in silico. TLR4-dependent viral entry was further validated by an anti-TLR4 antibody-mediated blocking experiment. It was noticed that TLR4 is necessary for the early events of viral infection, especially during the attachment and entry stages. Interestingly, it was also observed that TLR4 is not involved in the post-entry stages of CHIKV infection in host macrophages. The administration of TAK-242 decreased CHIKV infection significantly by reducing disease manifestations, improving survivability (around 75%) and reducing inflammation in mice model. Collectively, for the first time, this study reports TLR4 as one of the novel receptors to facilitate the attachment and entry of CHIKV in host macrophages, the TLR4-CHIKV-E2 interactions are essential for efficient viral entry and modulation of infection-induced pro-inflammatory responses in host macrophages, which might have translational implication for designing future therapeutics to regulate the CHIKV infection