Biology subjects

Savan, R.

Publications and source records attributed to Savan, R..

14 recordsLinked to original sources

bioRxiv2026

Myeloid IRF5 is required for TLR7-driven inflammatory hemophagocyte differentiation and Macrophage Activation Syndrome

In TLR7-driven macrophage activation syndrome (MAS), inflammatory hemophagocytes (iHPCs) differentiate from Ly6CHI monocytes, phagocytose red blood cells and promote disease, including anemia and thrombocytopenia. We demonstrate here that IRF5 is required for iHPC differentiation and MAS in TLR7-overexpressing (TLR7.1) mice. Both constitutive and myeloid-specific Irf5 deletion reduced iHPCs and improved anemia, thrombocytopenia and survival. Furthermore, therapeutic inhibition of IRF5 ameliorated MAS features and reduced splenic and circulating iHPCs. While cell-intrinsic IRF5 expression was required for iHPC differentiation, it was not required for TLR7.1 Ly6CHI monocyte differentiation and monocyte transcriptional programs. We further show that the transcriptome and chromatin landscape changed dramatically as iHPCs differentiated from TLR7.1 Ly6CHI monocytes. Many transcriptional programs gained in iHPCs were enriched in genes associated with IRF5-binding accessible chromatin regions, including those associated with NF-kB signaling, cytokine and chemokine production, and complement activation. Our data suggest that IRF5 collaborates with other transcription factor families, including NF-kB, ETS and AP1 members, to regulate iHPC gene programs. Together, our findings demonstrate that expression of IRF5 in myeloid cells is critical for MAS, for iHPC differentiation, and acts broadly across iHPC-specific gene programs in TLR7-driven inflammation.

immunology↗

bioRxiv2026

Lineage-specific chromatin poising enforced by ETS-IRF composite elements determines the divergent interferon responses of plasmacytoid dendritic cells and epithelial cells

Plasmacytoid dendritic cells (pDCs) produce robust type I interferons (IFN-I) within hours of viral sensing, while epithelial cells at mucosal surfaces mount a delayed response dominated by type III interferons (IFN-III). Both cell types express pattern recognition receptors that activate similar downstream transcription factors, yet they produce distinct subsets of IFNs. The mechanisms underlying these differences have remained unclear. Here, using Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) in primary human pDCs and intestinal epithelial cells, we show that IFN-I and IFN-III gene loci carry opposing, constitutively established chromatin accessibility landscapes that determine cell-type-specific interferon induction. The IFN-I locus is broadly open in pDCs and constitutively closed in epithelial cells, while the IFN-III locus displays the reciprocal pattern. Motif enrichment analysis of accessible regions at the IFN-I locus in pDCs revealed unexpected and significant enrichment of ETS family binding motifs alongside IRF motifs, which determines the cell-type-specific locus accessibility. The ETS factor PU.1 and IRF8 bind IFN-I promoters, at composite ETS-IRF elements positioned at the Positive Regulatory Domain IV (PRDIV) site of IFNB1 promoter and adjacent to the TATA-proximal IRF motif of IFNA promoters. IFNL gene promoters lack ETS recognition sequences. PU.1-IRF8 composite factor binding extends across intergenic regions of the IFN-I locus, where candidate enhancer elements marked by H3K4me1, H3K27ac, and RNA Pol II occupancy were identified. We propose that this network of ETS-IRF composite-element-anchored enhancers maintains the IFN-I locus in a constitutively poised state in pDCs, licensing the rapid and robust IFN-I response that defines pDCs. In epithelial cells, the absence of PU.1 and IRF8 renders the IFN-I locus epigenetically silent, while the IFN-III locus is constitutively open. Despite this, the delayed IFN-III gene expression in epithelial cells is due to intrinsically weaker promoter activity relative to IFN-I. These findings reveal that the divergent IFN induction of pDCs and epithelial cells is determined by the chromatin architecture prior to infection. Overall, these observations show that lineage-specific ETS-IRF regulatory factors and promoter strength determine the cell-type-specific IFN activation.

immunology↗

bioRxiv2025

Plasmacytoid dendritic cells in the intestine preferentially produce interferon lambda at homeostasis contributing to tonic localized innate immune responses

The healthy intestine maintains homeostasis in part via immune responses to microbiota, which includes basal production of interferon cytokines. Previous work showed that Type III Interferon (IFN-{lambda}) stimulates localized pockets of interferon-stimulated genes (ISGs) in the adult mouse intestinal epithelium at homeostasis that provide preemptive protection from viral pathogens. Here, we demonstrate that a major source of homeostatic IFN-{lambda} production in the intestine is a population of epithelium-associated plasmacytoid dendritic cells (pDC). Depletion of bacterial microbiota in the intestine also reduces pDC abundance, and pDC depletion or bone marrow reconstitution with IFN-{lambda}-deficient pDC results in reduced expression of homeostatic ISGs in the intestinal epithelium. Notably, intestinal pDC preferentially produce IFN-{lambda} over Type I IFNs whereas splenic pDC produce more Type I IFNs. Comparison of intestinal and splenic pDC reveal tissue-specific changes in gene expression and genomic accessibility, including evidence of responses to transforming growth factor beta (TGF-{beta}) in the intestine. Isolated gut pDC produce more IFN-{lambda} than splenic pDC upon stimulation, and pre-treatment of a human pDC cell line with TGF-{beta} results in enhanced production of IFN-{lambda} upon stimulation. This study demonstrates that pDC are an important source of homeostatic IFN-{lambda} in the intestine and defines the role of barrier cytokine TGF-{beta} in regulating IFN types produced by pDC upon stimulation. Reprogramming of recruited pDC by tissue cytokines may have important implications for balancing effective antimicrobial responses with damaging inflammation at barrier tissues. One sentence summaryThis study demonstrates that pDC are an important source of homeostatic IFN-{lambda} in the intestine and defines the role of barrier cytokine TGF-{beta} in regulating IFN types produced by pDC upon stimulation.

immunology↗

bioRxiv2025

Identification of host lncRNAs that impact Venezuelan equine encephalitis virus replication

Venezuelan equine encephalitis virus (VEEV) causes encephalitis in humans and equids, and there are no vaccines or therapeutics available for humans. In recent years, non-coding RNAs have emerged as critical regulatory factors affecting different cellular pathways. Specifically, long non-coding RNAs (lncRNAs) have been identified as regulators of antiviral pathways during various viral infections; however, their role in regulating VEEV infection has not been assessed. Here we show differential expression of several lncRNAs in primary mouse target cells infected with a vaccine strain of VEEV (TC-83) but not a pathogenic strain (TrD). Among the differentially expressed genes (DEGs), suppressing lncRNA small nucleolar RNA host gene 15 (Snhg15) resulted in about a 7-fold increase in VEEV TC-83 replication in primary mouse astrocytes. Knockdown of Snhg15 during VEEV TC-83 infection resulted in the suppression of ten genes including Irf1, Junb, Atf3, Relb, Pim1, Hbegf, Ccl5, Ankrd33b, and H2-K2, all of which were also increased during TC-83 infection when the expression of Snhg15 increased in primary mouse astrocytes. Most of these genes are involved in antiviral responses. KEGG pathway analysis confirmed the suppression of both pattern recognition receptor and inflammatory pathways after in Snhg15 knockdown. These data are the first to identify lncRNA responses in encephalitic alphavirus infection and demonstrate important roles for these overlooked RNAs on VEEV infection. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=186 SRC="FIGDIR/small/653438v1_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@15431eeorg.highwire.dtl.DTLVardef@9a22caorg.highwire.dtl.DTLVardef@1622310org.highwire.dtl.DTLVardef@1ebfd7b_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2025

Comprehensive Atomic-Scale 3D Viral-Host Protein Interactomes Enable Dissection of Key Mechanisms and Evolutionary Processes Underlying Viral Pathogenesis

Viral-human protein interactions are critical for viral replication and modulation of the host immune response. Structural modeling of these interactions is vital for developing effective antiviral therapies and vaccines. However, 99% of experimentally determined binary host-viral interactions currently lack structural information. We aimed to address this gap by leveraging computational protein structure prediction methods. Using extensive benchmarking, we found AlphaFold to be the most accurate structure prediction model for host-pathogen protein interactions. We then predicted the structures of 11,666 binary protein interactions across 33 viral families and created the most comprehensive atomic-scale 3D viral-host protein interactomes till date (https://3d-viralhuman.yulab.org). By integrating these interactomes with genetic variation data, we identified population-specific signatures of selection on variants coding for interfaces of viral-human interactions. We also found that viral interaction interfaces were less conserved than non-interface regions, a striking trend that is opposite to what is observed for host interfaces, suggesting different evolutionary pressures. Systematic analyses of interface sharing between host and viral proteins binding to the same host protein revealed mutation rate-dependent differences in interface mimicry. Similar mutation rate-dependent differences were seen in the interface sharing between viral proteins binding to a host protein. We also found that the patterns of E6 protein binding to KPNA2 differed between high- and low-risk oncogenic human papillomaviruses (HPVs), and clustering based on these binding patterns allowed the classification of HPVs with unknown oncogenic risk. Our interface mimicry analyses also unveiled a novel mechanism by which herpes simplex virus-1 UL37 suppresses the antiviral immune response through disruption of the TRAF6-MAVS signalosome interaction. Overall, our comprehensive 3D viral interactomes provide a resource at unprecedented scale and resolution that will enable researchers to explore how variation and signatures of selection influence viral interactions and disease progression. This tool also facilitates the identification of conserved and unique interaction patterns across viruses, empowering researchers to generate testable hypotheses and ultimately accelerate the discovery of novel therapeutic targets and intervention strategies.

systems biology↗

bioRxiv2025

CELF2 suppresses endogenous RNA ligands that activate RIG-I-mediated interferon induction

Type I interferons (IFNs) are critical for the control of viral infections, but aberrant IFN expression can result in tissue damage. RIG-I-like receptors (RLRs), such as RIG-I and MDA5, sense viral RNA and signal through the adaptor protein MAVS to induce phosphorylation and nuclear translocation of the transcription factor IRF3, thereby driving IFN production. However, activation of RLRs by endogenous RNA ligands can also induce IFNs, leading to autoinflammation. Identifying factors that suppress endogenous RNA ligands is critical for preventing IFN-induced autoimmunity. In this study, we identify a novel regulator, CELF2, that suppresses endogenous RNAs that otherwise activate the RLR pathway. We uncovered a novel role for the splicing factor CELF2 as a suppressor of immunostimulatory endogenous RNA ligands. Depletion of CELF2 in macrophages led to a spontaneous IFN and IFN-stimulated gene signature, dependent on the RIG-I-MAVS pathway. Furthermore, the transfer of RNA from CELF2-depleted macrophages was sufficient to induce type I IFN expression in naive cells. This RNA was found to be double-stranded as RNase III treatment of RNA derived from CELF2-depleted cells ablated IFN induction in naive cells. Immunoprecipitation of double-stranded RNA from CELF2-depleted macrophages revealed several immunostimulatory RNAs, which contribute to the increased interferon-stimulated gene signature observed in CELF2-depleted macrophages. These data indicate that CELF2 suppresses endogenous RNA ligands, which could otherwise activate RIG-I and induce an IFN signature. Overall, these findings reveal that CELF2 is an important regulator of self-RNA ligands to prevent IFN-induced autoinflammation. One-sentence summaryCELF2 suppresses RIG-I-like receptor ligands that activate interferon.

immunology↗

bioRxiv2023

Cellular RNA interacts with MAVS to promote antiviral signaling

Immune signaling needs to be well-regulated to promote clearance of pathogens, while preventing aberrant inflammation. Interferons (IFNs) and antiviral genes are activated by the detection of viral RNA by RIG-I-like receptors (RLRs). Signal transduction downstream of RLRs proceeds through a multi-protein complex organized around the central adaptor protein MAVS. Recent work has shown that protein complex function can be modulated by RNA molecules providing allosteric regulation or acting as molecular guides or scaffolds. Thus, we hypothesized that RNA plays a role in organizing MAVS signaling platforms. Here, we show that MAVS, through its central intrinsically disordered domain, directly interacts with the 3' untranslated regions of cellular mRNAs. Importantly, elimination of RNA by RNase treatment disrupts the MAVS signalosome, including newly identified regulators of RLR signaling, and inhibits phosphorylation of the transcription factor IRF3. This supports the hypothesis that RNA molecules scaffold proteins in the MAVS signalosome to induce IFNs. Together, this work uncovers a function for cellular RNA in promoting signaling through MAVS and highlights a generalizable principle of RNA regulatory control of cytoplasmic immune signaling complexes.

immunology↗

bioRxiv2022

Host specific sensing of coronaviruses and picornaviruses by the CARD8 inflammasome

Hosts have evolved diverse strategies to respond to microbial infections, including the detection of pathogen-encoded proteases by inflammasome-forming sensors such as NLRP1 and CARD8. Here, we find that the 3CL protease (3CLpro) encoded by diverse coronaviruses, including SARS-CoV-2, cleaves a rapidly evolving region of human CARD8 and activates a robust inflammasome response. CARD8 is required for cell death and the release of pro-inflammatory cytokines during SARS-CoV-2 infection. We further find that natural variation alters CARD8 sensing of 3CLpro, including 3CLpro-mediated antagonism rather than activation of megabat CARD8. Likewise, we find that a single nucleotide polymorphism (SNP) in humans reduces CARD8s ability to sense coronavirus 3CLpros, and instead enables sensing of 3C proteases (3Cpro) from select picornaviruses. Our findings demonstrate that CARD8 is a broad sensor of viral protease activities and suggests that CARD8 diversity contributes to inter- and intra-species variation in inflammasome-mediated viral sensing and immunopathology.

immunology↗

bioRxiv2022

MEF2A suppresses replicative stress responses that trigger DDX41-dependent IFN production

Interferons (IFN) are induced by sensing of self- and non-self DNA or genomic lesions by pathogen recognition receptors (PRR) that activate STING. These pathways must be kept in check by negative regulators to prevent unscheduled activation of IFN, which contributes to autoinflammation. Here we show that MEF2A as a novel negative regulator of inflammation that suppresses homeostatic induction of IFNs. Indeed, MEF2A deficiency results in the spontaneous production of type I IFN and robust downstream IFN-stimulated gene expression that coincided with a robust cellular antiviral state. Mechanistically, MEF2A depletion promoted the accumulation of R-loops that activate the DDX41/STING pathway. This pro-inflammatory pathway was dependent on ATR kinase activity, hallmark of the replicative stress response, was necessary for the activation of STING upon loss of MEF2A expression. Thus, our study connects MEF2A with protection from maladaptive type I IFN responses triggered by R-loop accumulation and links the DDX41-dependent activation of STING to the DNA damage response.

immunology↗

bioRxiv2022

Hemodynamic stress activates inflammatory responses and cell death through spectrin-dependent modulation of Store Operated Calcium Entry

Patients undergoing cardiac surgery face significant inflammatory induced by exposure to cardiopulmonary bypass (CPB), contributing to heightened morbidity and mortality. The molecular and cellular mechanisms that underpin this inflammatory process remain unknown. To address this knowledge gap, we performed snRNA/ATAC-Sequencing on leukocytes from neonatal CPB patients. Classical monocytes become more prevalent and have dysregulation of inflammatory genes after CPB, indicating their role in CPB-associated inflammation. A genome-wide CRISPR screen and in vitro experiments in non-adherent monocytic cells identified two novel genes, SPTAN1 and RAF1, as effectors of hemodynamic stress. SPTAN1 and RAF1 activate store-operated calcium entry that results inflammation and cell death. snATAC-Seq revealed dynamically changing patterns of chromatin accessibility and AP-1 transcription factor binding after CPB exposure. These findings provide novel insights into the pathogenesis of CPB-associated inflammation, with broad implications for understanding the early stages of sterile inflammation and how non-adherent cells sense shear stress.

cell biology↗

bioRxiv2021

Sequence diversity in the 3' untranslated region of alphavirus modulates IFIT2-dependent restriction in a cell type-dependent manner

Alphaviruses (family Togaviridae) are a diverse group of positive-sense RNA (+ssRNA) viruses that are transmitted by arthropods and are the causative agent of several significant human and veterinary diseases. Interferon (IFN)-induced proteins with tetratricopeptide repeats (IFITs) are a family of RNA-binding IFN stimulated genes (ISGs) that are highly upregulated following viral infection, and have been identified as potential restrictors of alphaviruses. The mechanism by which IFIT1 restricts RNA viruses is dependent on self and non-self-discrimination of RNA, and alphaviruses evade this recognition via their 5UTR. However, the role of IFIT2 during alphavirus replication and the mechanism of viral replication inhibition is unclear. In this study, we identify IFIT2 as a restriction factor for Venezuelan equine encephalitis virus (VEEV) and show that IFIT2 binds the 3 untranslated region (3UTR) of the virus. We investigated the potential role of variability in the 3UTR of the virus affecting IFIT2 antiviral activity by studying infection with VEEV. Comparison of recombinant VEEV clones containing 3UTR sequences derived from epizootic and enzootic isolates exhibited differential sensitivity to IFIT2 restriction in vitro infection studies, suggesting that the alphavirus 3UTR sequence may function in part to evade IFIT2 restriction. In vitro binding assays demonstrate that IFIT2 binds to the VEEV 3UTR, however in contrast to previous studies VEEV restriction did not appear to be dependent on the ability of IFIT2 to inhibit translation of viral RNA, suggesting a novel mechanism of IFIT2 restriction. Our study demonstrates that IFIT2 is a restriction factor for alphaviruses and variability in the 3UTR of VEEV can modulate viral restriction by IFIT2. Ongoing studies are exploring the biological consequences of IFIT2-VEEV RNA interaction in viral pathogenesis and defining sequence and structural features of RNAs that regulate IFIT2 recognition.

microbiology↗

bioRxiv2021

Active Translation Control of CD4 T Cell Activation by Regulatory T Cells

Increased protein synthesis is a hallmark of lymphocyte activation. Regulatory T cells (Tregs) suppress the activation and subsequent effector functions of CD4 effector T cells (Teffs). However, molecular mechanisms that enforce Treg-mediated suppression in CD4 Teff are not fully clear. Control of CD4 Teff activation by Tregs has largely been defined at the transcriptional level, which does not reflect changes in post-transcriptional control. We found that Tregs suppressed activation-induced global protein synthesis in CD4 Teffs prior to cell division. We analyzed genome-wide changes in the transcriptome and translatome of activated CD4 Teffs using two independent approaches. We show that mRNAs encoding for the protein synthesis machinery are regulated at the level of translation in activated Teffs. Strikingly, Tregs suppressed global protein synthesis of CD4 Teffs by specifically inhibiting mRNAs of the translation machinery at the level of mTORC1-mediated translation control. Lastly, we found that the RNA helicase eIF4A inhibitor rocaglamide A (RocA) can suppress CD4 Teff activation in vitro to alleviate inflammatory CD4 Teff activation caused by acute Treg depletion in vivo. These data provide evidence that peripheral tolerance is enforced by Tregs through mRNA translational control in CD4 Teffs. Therefore, therapeutic targeting of the protein synthesis machinery can be expected to mitigate inflammatory responses invoked by Treg loss of function.

immunology↗

bioRxiv2021

Endomembrane targeting of human OAS1 p46 augments antiviral activity

Many host RNA sensors are positioned in the cytosol to detect viral RNA during infection. However, most positive-strand RNA viruses replicate within a modified organelle co-opted from intracellular membranes of the endomembrane system, which shields viral products from host cell innate immune sensors. Targeting innate RNA sensors to the endomembrane system may enhance their ability to sense viral RNA generated by viruses that use these compartments for replication. Here, we reveal that an isoform of oligoadenylate synthetase 1, OAS1 p46, is prenylated and targeted to the endomembrane system. Membrane localization of OAS1 p46 confers enhanced access to viral replication sites and results in increased antiviral activity against a subset of RNA viruses including flavivirus, picornavirus, and SARS-CoV-2. Finally, our human genetic analysis shows that the OAS1 splice-site SNP responsible for production of the OAS1 p46 isoform strongly associates with COVID-19 severity. This study highlights the importance of endomembrane targeting for the antiviral specificity of OAS1 and suggests early control of SARS-CoV-2 replication through OAS1-p46 is an important determinant of COVID-19 severity.

immunology↗

bioRxiv2019

A sustained small increase in NOD1 expression promotes ligand-independent oncogenic activity

Small genetically-determined differences in transcription (eQTLs) are implicated in complex disease but the mechanisms by which small changes in gene expression impact complex disease are unknown. Here we show that a persistent small increase in expression of the innate sensor NOD1 precipitates large cancer-promoting changes in cell state. A ~1.2-1.4 fold increase in NOD1 protein concentration by loss of miR-15b/16 regulation sensitizes cells to ligand-induced inflammation, with an additional slight increase leading to ligand-independent NOD1 activation that is linked to poor prognosis in gastric cancer. Our data show that tight expression regulation of NOD1 prevents this sensor from exceeding a physiological switching checkpoint that promotes persistent inflammation and oncogene expression and reveal the impact of a single small quantitative change in cell state on cancer.\n\nOne Sentence SummaryA small change in NOD1 expression has a large cancer-promoting impact on cell state.

systems biology↗