Soma Chattopadhyay is funded by the Department of Science and Technology (DST), New Delhi, India, vide\grant no EMR/2016/000854. the data suggest that the viral PAMPs\induced IFN response is usually negatively regulated by IRGM. Several classical cell\autonomous viral restriction factors are upregulated in IRGM\depleted cells Viral restriction factors are host cellular proteins that establish the first line of defense and are capable of blocking almost all stages of the viral life cycle, including viral entry, replication, assembly, and egress/release (Goff, 2004; Colomer\Lluch and and and and and and and family. We found that CHIKV replication was significantly reduced (80C90%, 24 hpi) in IRGM\deficient cells (Fig?3A). Consistently, there was a strong reduction (10 fold) in total infectious computer virus particles produced by IRGM knockdown cells compared to the control in plaque assays (Fig?3B and C). Further, in agreement, replication of CHIKV was found to be significantly higher in IRGM overexpressing cells (18 hpi) (Fig?EV3A). Open in a separate FPS-ZM1 window Physique 3 IRGM\depleted cells can restrict contamination with CHIKV, HSV\1, JEV, VSV, ZIKV, WNV, and SARS\CoV\2 Total RNA was isolated from mock and CHIKV (MOI 1, 24?h) infected control and IRGM knockdown HT\29 cells and subjected to qRTCPCR with CHIKV specific primers to quantitate total viral load (family. We observed a significant inhibition (??90% at 24 hpi) of FPS-ZM1 HSV\1 replication in IRGM\depleted HT\29 cells compared to the control cells (Fig?3D). Measurement of produced infectious HSV\1 particles using the plaque assay also showed a dramatic reduction in FPS-ZM1 IRGM\deficient cells in comparison to the controls (Fig?3E and F). We next tested the replication of three positive\sense ssRNA viruses of the family, JEV, ZIKV, and WNV. As compared to control cells, IRGM knockdown cells showed inhibition of JEV replication and infectious particle egression of TNFRSF1B ?50% and 90%, respectively (Fig?3GCI). The replication of the ZIKV and WNV was compared in control and IRGM\depleted Huh7 cells. The percentage of infected cells FPS-ZM1 was determined by flow cytometry after staining cells with primary antibody against protein E, a flavivirus group antigen. This analysis revealed that IRGM\depleted cells could robustly restrict the growth of the ZIKV and WNV (Fig?3J and M). In plaque assays, more than 90% inhibition of WNV (Fig?3K and L) and ZIKV computer virus was observed in IRGM knockdown cells (Fig?3N and O). We also tested replication of VSV, a negative sense ssRNA computer virus from the family that mainly infects animals and is the most common computer virus used in the laboratory to study immune responses. Strong inhibition of VSV replication was observed in IRGM knockdown HeLa cells compared to the control cells (Fig?EV3B). We uncovered the control and IRGM knockdown HeLa and THP\1 cells to a VSV strain expressing GFP for 4?h. The GFP fluorescence was first observed in control cells as compared to IRGM knockdown cells and also a lower fluorescence was observed in IRGM knockdown HeLa and THP\1 cells (Fig?EV3C) indicating a reduced propagation of the computer virus. Further, we tested whether the complementation of IRGM in IRGM knockdown cells can rescue the viral replication defect. Indeed, the viral replication in Flag IRGM complemented IRGM knockdown cells was similar to wild\type cells (Fig?EV3D). The ongoing SARS\CoV\2 outbreak has posed an enormous threat to global public health. SARS\CoV\2 is usually a positive\sense ssRNA computer virus belonging to the family. A strong induction of type I IFN response upon SARS\CoV\2 contamination is usually reported (Winkler and and and.