bluuebunny/crossref_metadata_2025_split huggingface dataset:
The E3-ligase TRIM7 acts as an antiviral factor by ubiquitinating the SARS-CoV-2 membrane protein to limit apoptosis and virus replication
Authors: Gonzalez Orozco Maria J , Hage Adam , Xia Hongjie , Huante Matthew , van Tol Sarah , Giraldo Maria I , Puebla-Clark Lucinda , Menachery Vineet , Stephens Robin , Endsley Mark , Endsley Janice , Shi Pei-Yong , Freiberg Alexander , Rajsbaum Ricardo 
Published: January 2023

Abstract
Cytokines, such as type-I Interferons (IFN-I), are essential to mount an effective antiviral response. The production of IFN-I is regulated by multiple mechanisms, including ubiquitination of host signaling factors. However, viruses can hijack the ubiquitin (Ub) system to promote viral replication. For example, we recently reported that the E3-Ub ligase TRIM7 ubiquitinates the envelope protein of Zika virus, which increases virus binding to its receptor favoring virus entry. Previous studies suggest that TRIM7 may play dual roles during infection by promoting or inhibiting IFN-I induction depending on the conditions. Here, we used our recently generated Trim7−/− mice to study its physiological role during SARS-CoV-2 (SCoV2) infection. We found that Trim7−/− mice have increased weight loss and lung virus titers compared to WT controls upon infection with SCoV2. We also found a dysregulated innate immune response, which included increased IFN-I induction and reduced neutrophil recruitment and production of inflammatory cytokines such as IL-6 in Trim7−/− mice. We observed a higher frequency of apoptotic cells in Trim7−/− compared to WT mice, specifically in CD45− lung epithelial cells. Mechanistically, using co-immunoprecipitation studies, we confirmed previous reports that TRIM7 interacts with the viral membrane protein (M), which can cause apoptosis. Using recombinant mutants that lack ubiquitination sites on M, we found that TRIM7-mediated ubiquitination of M protects cells against death. This suggests that TRIM7 restricts SCoV2 infection via a novel non-degradative mechanism involving ubiquitination of M. Understanding the role of TRIM7 in inhibiting SCoV2 replication could help develop treatments for COVID-19.
Supported by grants from NIH R01 AI166668
Differential regulation of macrophage galactose-type lectin (MGL) orthologs by mycobacterial ligands
Authors: Paez Reina N , Browning Sarah , Files Megan , Varghese Alyssa , Naqvi Kubra F , Huante Matthew B , Strong Emily , Lee Sunhee , Endsley Mark , Endsley Janice J 
Published: January 2023

Abstract
Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB), is responsible for approximately 10 million new infections and 1.5 million deaths each year. Pattern recognition receptors (PRRs) expressed on the surface of macrophages play an important role in orchestrating the immune response to pathogens. Amongst these, the C-type lectin receptors (CLRs) are an important PRR system that is increasingly understood to mediate innate immunity to mycobacteria. We recently described a new anti-bacterial and anti-inflammatory role for the macrophage galactose-type lectin-1 (MGL-1) in a murine model of experimental TB. MGL is a type II CLR that is known to recognize galactose and N-acetylgalactosamine monosaccharides. We observed that exposure to Mtb activated expression of both murine MGL orthologues, MGL-1 and MGL-2. The specific pathogen-associated molecular patterns that activate MGL orthologue transcription, however, have yet to be determined. Here we demonstrate that expression of MGL-1 and MGL-2 are differentially regulated by cytokines, TLR agonists, and mycobacterial ligands. MGL-1 expression is activated by dexamethasone, IL-4, IL-10, and the mycobacterial cell wall glycolipid Trehalose 6,6′-dimycolate. In contrast, MGL-2 expression is activated by TLR2/6 and TLR2/1 agonists and total lipid extracts of mycobacteria. Use of a MyD88 inhibitor further showed that molecular regulation of MGL-1 and MGL-2 transcription, following exposure of macrophages to mycobacteria, is TLR pathway-dependent. These results identify overlapping and unique host immune pathways that differentially activate MGL orthologues following exposure to Mtb and could be targeted through immune modulation.
Recombinant Fasciola hepatica fatty acid binding protein (Fh15) as a novel anti-inflammatory biotherapeutic in an acute gram-negative non-human primate sepsis model
Authors: Espino Ana M , Rosado-Franco Jose J , Armina-Rodriguez Albersy , Marzan-Rivera Nicole , Burgos Armando G , Spiliopoulus Natalie , Mendez Loyda B 
Published: January 2023

Abstract
Due to their phylogenetic proximity to human, non-human primates are considered an adequate choice for basic and pre-clinical model of sepsis. Gram-negative bacteria are the primary causative of sepsis. During infection, bacteria continuously release the potent toxin lipopolysaccharide (LPS) into the bloodstream, which triggers an uncontrolled systemic inflammatory response leading to death. Our previous research has demonstrated in vitro and in vivo using a mouse model of septic shock that Fh15, a recombinant variant of the Fasciola hepatica fatty acid binding protein, acts as an antagonist of TLR4 suppressing the LPS-induced pro-inflammatory cytokine storm. The present communication is a proof-of concept study aimed to demonstrate that a low-dose of Fh15 suppresses the cytokine storm and other inflammatory markers during the early phase of sepsis induced in rhesus macaques by i.v. infusion with lethal doses of live E. coli. Fh15 was administrated as isotonic infusion 30 min prior to the bacterial infusion. Among the novel findings reported in this communication, I) Fh15 significantly prevented bacteremia, suppressed LPS levels in plasma and the production of C-reactive protein and procalcitonin, which are key signature of inflammation and bacterial infection, respectively, II) reduced the production of pro-inflammatory cytokines, and III) increased innate immune cell populations in blood, which suggest a role in promoting a prolonged steady state in rhesus macaques even in the presence of inflammatory stimuli. This is the first report demonstrating that a F. hepatica-derived molecule possesses potential as anti-inflammatory drug against sepsis in an NHP-model.
This research was supported by the National Institute of Allergy and Infectious Diseases Grant/Award Number: 1SC1AI155439-01, Office of Research Infrastructure Program of NIH (ORIP-NIH) 2016-2021, Grant/Award Number: P40OD12271; National Institute on Minorities Health and Health Disparities, Grant/Award Number: 5R25GM061151, G12MD007600 and R25GM061838.
Glycolysis is critical for granulocytic myeloid-derived suppressor cell (G-MDSC) activity during Staphylococcus aureus biofilm infection
Authors: Horn Christopher Michael , Heim Cortney E , Kielian Tammy L 
Published: January 2023

Abstract
Staphylococcus aureus (S. aureus) is a leading cause of prosthetic joint infection (PJI). These infections are often intractable due to biofilm formation, which are complex bacterial communities that adhere to biotic and abiotic surfaces. Within a biofilm, there is considerable heterogeneity in bacterial metabolism and gene expression that serves to create an anti-inflammatory milieu at the site of infection. This is due, in part, to the preferential recruitment of granulocytic myeloid-derived suppressor cells (G-MDSCs), a population of pathologically activated immature leukocytes with immunosuppressive features. Our prior work revealed that G-MDSCs are integral for promoting biofilm persistence via the production of interleukin-10 (IL-10) in response to biofilm-derived lactate. However, the mechanism responsible for G-MDSC anti-inflammatory activity in the context of biofilm infection is unknown. Using a mouse model of S. aureus PJI, time-course single-cell RNA sequencing (scRNA-seq) revealed a significant enrichment in glycolytic and hypoxic transcriptional signatures in infiltrating G-MDSCs. G-MDSCs significantly increased their glycolytic activity when co-cultured with S. aureus biofilm in vitro and inhibiting glycolysis during PJI in vivo with 2-DG nanoparticles significantly attenuated G-MDSC suppressive activity concomitant with a significant reduction in bacterial burden. This study demonstrates that glycolysis is required for G-MDSC-mediated immunosuppression and biofilm persistence during S. aureus PJI. Furthermore, our results demonstrate that G-MDSC metabolism can be targeted to improve infection outcomes.
Supported by funding from NIH (2P01 AI083211) & UNMC (UNMC Graduate Fellowship)
Identifying innate immune receptors causing production of inflammatory cytokines by infection of Francisella tularensis.
Authors: Gutierrez Stephanie K , Spencer Charles T , Setzu Nicole 
Published: January 2023

Abstract
Francisella tularensis is a pathogenic species and a causative agent of tularemia. This bacterium is incredibly infectious and often lethal in that F. tularensis induces the overproduction of pro-inflammatory cytokines. This study aims to identify the innate immune receptors responsible for the production of inflammatory cytokines in response to F. tularensis infection. The study uses an immortalized C57BL/6 Bone Marrow Macrophage (BMMac) cell line as the control and knockout BMMac deficient in various innate immune receptors. The cells are passaged and seeded before infecting them with F. tularensis for 2 hours. The cells were treated with a high dose of gentamicin for 2 hours, followed by a low dose of gentamicin for an incubation period of 24 hours. The supernatant from these cells is then collected and analysis processed by ELISA, an enzyme-linked immunosorbent assay, to measure the production of inflammatory cytokines. The results were compared with the positive control of wild-type BMMac and the negative control of uninfected BMMac cells. Analysis of these mutants reveals the involvement of innate immune receptors in triggering the pro-inflammatory cytokine storm in response to F. tularensis infection that leads to the host’s death.
Supported by NIH (SC1 GM122699-01A1)
Merocytophagy contributes to dissemination of Francisella tularensis and is enhanced by SYK signaling in macrophages
Authors: Deobald Kelly N , Steele Shaun P , Kawula Thomas H 
Published: January 2023

Abstract
Merocytophagy is a phagocytic process characterized by the transfer of cytosolic content between macrophages and the formation of a double-membraned vesical within recipient cells. Studies conducted in our laboratory have demonstrated that proteins and intracellular pathogens can be transferred between cells by this contact-dependent mechanism. In particular, Francisella tularensis, a highly infectious pathogen that spreads from cell to cell by hijacking the phagocytic pathway in macrophages, has been shown to replicate within cells that receive cargo via merocytophagy. While these observations may point to an alternative route of F. tularensis dissemination in vivo, it is also possible that this mechanism of bacterial spread could amplify immune responses. Ongoing studies aim to characterize molecular and cellular players in merocytophagy to gain a better understanding of this phenomenon and to develop strategies to assess merocytophagy in vivo in the future. In cell-to-cell contact studies, inhibition of spleen tyrosine kinase (SYK) expression and activity showed decreased cargo transfer by merocytophagy. Pharmacologic inhibition of SYK was also associated with decreased surface expression of several integrins, giving evidence that cell-to-cell contact is stabilized by integrins during merocytophagic transfer in macrophages.
Supported by grants from NIH and NIGMS (R56 AI139476 and T32 GM008336).
UBX Domain Protein 6 Positively Regulates JAK-STAT1/2 Signaling
Authors: Harrison Andrew G , Ketkar Harshada , Wang Penghua 
Published: January 2023

Abstract
Type I/III IFNs induce expression of hundreds of IFN-stimulated genes through the JAK/STAT pathway to combat viral infections. Although JAK/STAT signaling is seemingly straightforward, it is nevertheless subjected to complex cellular regulation. In this study, we show that an ubiquitination regulatory X (UBX) domain-containing protein, UBXN6, positively regulates JAK-STAT1/2 signaling. Overexpression of UBXN6 enhanced type I/III IFNs–induced expression of IFN-stimulated genes, whereas deletion of UBXN6 inhibited their expression. RNA viral replication was increased in human UBXN6-deficient cells, accompanied by a reduction in both type I/III IFN expression, when compared with UBXN6-sufficient cells. Mechanistically, UBXN6 interacted with tyrosine kinase 2 (TYK2) and inhibited IFN-β–induced degradation of both TYK2 and type I IFNAR. These results suggest that UBXN6 maintains normal JAK-STAT1/2 signaling by stabilizing key signaling components during viral infection.
Supported by R01AI132526 and R21AI155820 to Penghua Wang
CD138 contributes to plasma cell dynamics and competition in the bone marrow
Authors: Park Rosa , Benet Zachary , Fooksman David R 
Published: January 2023

Abstract
Plasma cells (PCs) are terminally-differentiated, non-proliferative immune cells that mainly reside in the bone marrow (BM) and specialize in antibody secretion. Previous work in the laboratory has shown that CD138, the canonical PC surface marker, mediates PC survival in a cell-intrinsic manner and is required for long-term humoral immunity. Since CD138 surface expression levels are heterogeneous among PCs, we hypothesized that they regulate PC function and survival. We co-transferred WT and CD138+/− B cells into WT or CD138+/− recipient mice, immunized them, and tracked WT versus CD138+/− PCs. Consistent with their genotypes, donor-derived WT PCs express higher CD138 levels than CD138+/− PCs. In WT recipient BM, donor-derived WT PCs outnumber CD138+/− PCs. Yet in CD138+/− recipient BM, donor-derived WT and CD138+/− PC frequencies are similar, suggesting that PC competition affects survival in the BM and is mediated by CD138 expression. Our previous intravital imaging studies had shown that BM PCs are motile and organize into clusters, and these behaviors are regulated by adhesion molecules and cytokines. Since CD138 interacts with an array of adhesion ligands and cytokines, CD138 may enable PCs to efficiently scavenge for survival factors and assemble into clusters that may control survival or function. To test this idea, we compared movement and cluster formation among WT, CD138+/−, and CD138−/− PCs in the BM using intravital imaging. We also characterized surface marker and gene expression profiles of WT, CD138+/−, and CD138−/− PCs to gain further insight into how CD138 levels affect PC function. These studies will improve our understanding of factors governing PC function and survival, with implications for vaccine development.
Supported by grants from NIH (R01 HL141491, T32 GM007288)
SHIP-2 inhibits human microglia-like cell function in a TREM2 independent manner
Authors: Ramakrishnan Gautham S , Humphrey Mary Beth 
Published: January 2023

Abstract
Microglia are a crucial component of the immune response in neurodegenerative diseases including Alzheimer’s Disease (AD). The Triggering Receptor Expressed on Myeloid Cells #2 (TREM2) mediates microglial phagocytosis and migration, and promotes metabolic homeostasis. Src homology domain-containing inositol 5-phosphatase 1 (SHIP-1) inhibits TREM2 in myeloid cells like macrophages and osteoclasts. SHIP-2, a paralog of SHIP-1, inhibits other myeloid cell receptors including Macrophage Colony-Stimulating Factor (M-CSF). We hypothesize that SHIP-2, inhibits TREM2-mediated microglial functions. To test our hypothesis, we studied the effect of pharmacological inhibition of SHIP-2 (and SHIP-1) in the function of wild-type (HMC3WT) and Trem2−/− human microglia-like cells (HMC3T2KO), which predominantly express SHIP-2 compared to SHIP-1. HMC3WT displayed superior phagocytic ability of oligomerized amyloid β and apoptotic neurons, have a larger lysosomal compartment, and have increased mitochondrial activity compared to HMC3T2KO. Treatment with a pan SHIP-1/2 inhibitor (SHIPi) K161 significantly improved the phagocytic uptake of fluorescently tagged oligomerized Aβ42 in both HMC3WT and HMC3T2KO. K161 treatment resulted in an enlarged lysosomal compartment and increased mitochondrial activity in both HMC3WT and HMC3T2KO. Our data suggests that, in cases of low SHIP-1 expression, pan SHIP inhibition improves microglia-like cell functions independently of TREM2. Further studies to look for SHIP2-TREM2 interactions and the effect of SHIP-1 expression in combination with SHIPi treatment in HMC3 cell function are underway. SHIPi are non-toxic in HCM3 cells and may offer potential treatment options for AD.
Supported by grants from NIH (1RF1AG059717-01)
C3aR dampens intracellular cAMP to control TRIF-mediated inflammation in activated macrophages
Authors: Corcoran Jesse , Hickman James , Napier Brooke 
Published: January 2023

Abstract
Recent studies from our lab and others have shown that complement components can enhance cell-intrinsic inflammatory signaling pathways. Specifically, we have found that activation of complement receptor C3aR enhances TLR4 signaling in lipopolysaccharide (LPS)-treated macrophages via enhanced p-38 phosphorylation. We found C3aR−/− mice treated with systemic LPS have decreased inflammatory cytokines (IL-6 and TNF) compared to wild-type (WT) mice. Our results corroborate previous studies showing C3aR−/− mice display decreased systemic cytokines (IL-6 and TNF) and increased bacterial burden during Gram-negative (GN) infection compared to WT mice. These results suggest C3aR is a critical regulator of TLR4 signaling. TLR4-TRIF signaling leads to activation of IRF3 and expression of interferon-regulated genes. We find that primary bone marrow-derived macrophages (BMDMs) from C3aR−/− mice show significantly reduced TRIF-dependent ifnb expression and IFNb secretion following 24h LPS exposure. Additionally, WT BMDMs treated with a C3aR antagonist (C3aRi) show a similar reduction in ifnb expression after LPS-treatment. These data demonstrate C3aR is critical for TRIF-dependent signaling downstream of TLR4 activation. Here we find WT BMDMs treated with C3aRi and LPS show increased intracellular cAMP. These findings demonstrate C3aR activation leads to depleted cAMP, which is essential for TRIF-mediated signaling downstream of TLR4. We hypothesize that C3aR is working as an inhibitory GPCR to regulate inflammation via intracellular cAMP and enhancing TRIF signaling in LPS-treated macrophages. Our findings are the first to suggest C3aR reduces cAMP to control TRIF-mediated inflammation in activated macrophages.
IL-17 cytokines preferentially act on naive CD4+ T cells with IL-17AF inducing the greatest transcriptomic changes.
Authors: Crawford Michael Patrick , Borcherding Nicholas , Karandikar Nitin 
Published: January 2023

Abstract
CD4+ T-helper 17 (Th17) T cells are important in the regulation of protective immunity during infection and in self-tolerance/autoimmunity. Th17 cells, through the secretion of IL-17, act in promotion of inflammation and are thus a major potential therapeutic target in autoimmune disorders. Recent reports show that the IL-17 family cytokines, IL-17A, IL-17F and IL-17AF, can directly act on CD4+ T-cells, inducing functional changes in these cells. Here we show that functional and transcriptomic changes were preferentially induced in naïve CD4+ T-cells, but not in memory CD4+ T-cells. As early as 48 hours post-IL-17 exposure, naïve cells showed transcriptome changes, whereas memory cells remained unaffected as late as 7 days. The functional differences occurred despite similar IL-17RC and IL-17RA receptor expression on these subsets. Also, the differential IL-17-mediated changes in naïve, but not memory, cells occurred even in transwell/co-culture systems, with each subset maintaining its respective functional phenotype. Notably, there were differences in downstream transcriptional signaling by the three IL-17 cytokines. The IL-17AF heterodimer conferred both the greatest transcriptional change and most altered functional consequences. Detailed analysis of the transcriptome provides important insights into the genes and pathways that are altered as a result of IL-17-induced signaling and may serve as targets of future therapies.
This work was supported, in part, by grant awards from the NIH to N.J.K. (R01 AI121567) and N.B. (F30 CA29655).
Loss of the nuclear receptor PPARγ primes CD8+ T cells for exhaustion
Authors: Shah Rushil , Christofides Anthos , Aksoylar Halil-Ibrahim , Pal Rinku , Boussiotis Vassiliki A , Patsoukis Nikolaos 
Published: January 2023

Abstract
Peroxisome proliferator activated receptors (PPARs) are transcription factors that belong to nuclear hormone superfamily, with three distinct types identified: PPARapha (PPARα), PPARgamma (PPARγ), and PPARbeta/delta (PPARβ/δ). The role of PPARγ in the immune system has been mainly studied in macrophages, where it is known to mediate M2 polarization. Synthetic ligands for activating PPARγ ameliorate disease outcomes in various models of inflammation and autoimmunity. Here, we investigated how PPARγ affects antigen-specific effector and memory responses of CD8+ T cells using OT-I TCR transgenic mice and OT-I mice with T cell-specific PPARγ deletion. During CD8+ T effector differentiation in vitro by IL-2 culture following stimulation with cognate antigen, PPARγ deletion resulted in a rapid and enhanced T effector cell expansion and activation. However, PPARγ deficient OTI cells were more prone to exhaustion. This effect coincided with decreased mitochondrial fitness and spare respiratory capacity, high ROS levels and a significant increase in oxidized lipids. During generation of T effector cells in vivo, PPARγ deletion resulted in similar outcomes. At the memory phase there were no numerical differences in the T memory subsets in vivo, but upon rechallenge in vitro, PPARγ-deficient antigen-specific T cells had impaired capacity for effector differentiation. Our findings uncover a new role of PPARγ in CD8+ T responses and indicate that PPARγ has an indispensable role in regulating metabolism-mediated cues that imprint T cell differentiation and long-term effector capacity.
Supported by RO1CA212605, RO1CA229784-01, RO1CA238263 (to VAB)
Optimal CD8+ T cell effector function requires costimulation-induced RNA-binding proteins that reprogram the transcript isoform landscape
Authors: Karginov Timofey , Menoret Antoine , Vella Anthony T 
Published: January 2023

Abstract
Boosting T cell activation through costimulation directs defense against cancer and viral infections. Despite many studies targeting costimulation in clinical trials, the increased potency and reprogramming of T cells endowed by costimulation is poorly understood. Canonical dogma states that transcription mediates T cell activation. Here, we show that the spliceosome, controlling post-transcriptional alternative splicing and alternative polyadenylation, is the most significantly enriched pathway in T cells after CD134/CD137 costimulation. Costimulation of CD8+ T cells significantly increased expression of 29 RNA-binding proteins (RBPs) while RNA-seq uncovered over 1000 differential alternative splicing and polyadenylation events. Using in vivo mouse and in vitro human CRISPR models, we demonstrate that RBP Tardbp, among multiple other RBPs, is required for effector cytokine production, CD8+ T cell clonal expansion, and isoform regulation after costimulation. The prospect of immune response optimization through reprogramming of mRNA isoform production offered herein opens new avenues for experimentally and therapeutically tuning the activities of T cells.
Supported by NIH grants AI136955 and R21AI139891.