bluuebunny/crossref_metadata_2025_split huggingface dataset:
The dynein-binding protein DISC1 plays important roles in relocating organelles to the immunological synapse during T cell activation.
Authors: Poenie Martin F. , Maskalenko Nicholas A. , Nath Shubhankar 
Published: January 2023

Abstract
DISC1 (Disrupted in Schizophrenia-1) is a dynein-binding scaffold protein that interacts with many other proteins. In neurons, DISC1 is important for proliferation, migration, and signaling. We found that DISC1 is expressed in Jurkat cells and human NK cells and that it coimmunoprecipitates with dynein in Jurkat cells. Immunostaining for DISC1 in unstimulated Jurkat cells showed that it was generally located around the MTOC. When Jurkat cells were stimulated with SEE-coated Raji cells, depending on the antibody used, we found DISC1 either clustered around mitochondria or colocalized with dynein and NDE1 at the immunological synapse (IS). Biochemical studies showed that after Jurkat cells are activated (SEE-coated Raji cells or PMA), DISC1 coimmunoprecipitates with talin and paxillin. Sequence analysis of cloned DISC1 cDNAs from Jurkat cells revealed that at least two isoforms (L and Lv) are expressed. When GFP-DISC1 isoforms were individually expressed in Jurkat cells, we found that the Lv isoform concentrated around mitochondria whereas the L isoform colocalized with dynein at the synapse as seen in the immunostaining data. While RNAi was relatively ineffective at reducing DISC1 expression, we could abolish expression by using a DISC1-targeted CAS9/CRISPR construct. In these DISC1-disrupted cells, we found that mitochondria failed to accumulate at the immunological synapse and the MTOC did not appear to move all the way to the contact site (perhaps suggesting a defect in docking). When GFP-DISC1 Lv was expressed in the DISC1 CAS9/CRISPR cells mitochondria accumulation at the IS was restored. Similarly, expression of the GFP-DISC1 L restored the normal positioning of the MTOC upon T cell activation.
Transcriptional profiling of pathogen-specific CD4 T cells reveals key events that regulate Th17 cell priming and differentiation
Authors: Gao Yajing , Deason Krystin L. , Jain Aakanksha , Dozmorov Igor , Wakeland Edward K. , Pasare Chandrashekhar 
Published: January 2023

Abstract
CD4 T cells play a central role in adaptive immunity through secretion of specific cytokines which exert effector functions against invading pathogens. They also drive B cell activation and coordinate CD8 T cell responses. Productive CD4 T cell responses are determined not only by the specificity of the T cell receptor, but also by innate immune cues provided by dendritic cells (DCs) which shape the nature of the effector lineage commitment. However, to what extent pathogen-recognizing DCs prime naïve T cells on a global transcriptional level remains elusive, because of the inability to track and identify pathogen-specific T cells during cognate DC-T cell interaction. Here, we analyzed transcriptional profiles of pathogen-specific mouse Th17 cells utilizing an in vitro approach, where naïve CD4 T cells were differentiated by mature DCs in a highly antigen-specific manner. These pathogen-specific Th17 cells exhibit enormous lineage heterogeneity and resemble Th17 cells found in vivo following infections. We further discovered that DC-mediated priming not only generates a distinct transcriptional landscape in pathogen-specific Th17 cells, but also warrants their acquisition of memory gene signature and metabolic switch from glycolysis to oxidative phosphorylation. These results indicate that pathogen-activated dendritic cells play a critical role in CD4 T cell transcription programming that extends beyond determining lineage commitment. By mining this dataset, we have identified a previously unappreciated gene in promoting Th17 lineage commitment. Conceptually, our study underlines the necessity of investigating physiologically differentiated CD4 T cells in order to reveal novel regulators of T cell biology.
Role of TNF-α reception in epidermal γδ T cell function during homeostasis and after TCR stimulation
Authors: Matthews Carolyn J. , Barba Cindy , Mendoza German , Jameson Julie M. 
Published: January 2023

Abstract
Inflammatory responses mediated by the cellular components of epithelial tissues are essential to the wound healing process. Epidermal γδ T cells, also known as dendritic epidermal T cells (DETC), play key roles in the wound healing process by producing growth factors, chemokines and cytokines necessary for the proliferation and migration of epithelial cells. DETC activation occurs via the γδ TCR, however, second and third signals are required to sustain the activation and production of factors needed in functions such as wound repair. While progress has been made on elucidating signal 2 costimulatory signals such as JAML and CAR, less is understood about what signals are required for signal 3 in DETC and what roles TNF-α plays in regulating and sustaining DETC function. TNF-α signals through two receptors, tumor necrosis factor receptor 1 and 2 (TNFR1 and TNFR2). These receptors elicit varied responses that include apoptosis, survival or inflammation. Our data indicate that DETC upregulate type 1 chemokines and their corresponding chemokine receptor expression during inflammation. We have also found that a subset of DETC express TNFR1 and TNFR2 on their cell surface constitutively suggesting a requirement for tonic TNFR signaling for particular DETC populations. We have quantified chemokine, cytokine and growth factor production from DETC stimulated in the presence or absence of TNF-α using Luminex technology. Additionally, we have investigated which TNFR is responsible for modulating DETC function by comparing DETC function in cells isolated from wild-type, TNFR1 and/or 2 knockout mice. Identifying which factors regulate DETC function and activation could provide potential targets for improving DETC function in wound repair.
IRF4 regulates the rate of cell cycle during B cell differentiation.
Authors: Patterson Dillon , Scharer Christopher D. , Boss Jeremy M. 
Published: January 2023

Abstract
Cell division is required for the initiation of B cell differentiation, the regulation of isotype class switching, and ultimately entry of cells into the plasma cell (PC) lineage. Division coupled changes in the expression of transcription factors, such as interferon regulatory factor-4 (IRF4), that coordinate the PC transcriptional program also occur; however, little is known regarding how these factors coordinate cell division and exit from the cell cycle after differentiation. To begin to address this gap in knowledge, we assessed the cell division kinetics of wildtype (WT) B cells responding to the T cell independent antigen lipopolysaccharide (LPS). Interestingly, we found that WT B cells undergo at least 8 divisions before differentiating into PCs. Computational modeling of division rates over time defined a proliferative burst between 48 and 60 hours after LPS injection that is characterized by a rapid increase in the rate of cell division. IRF4-deficient B cells divided but failed to undergo the proliferative burst resulting in fewer activated B cells. Indeed, IRF4 is expressed at low levels during these divisions before being upregulated in post-mitotic CD138+ PCs after 8 divisions. Notably, while more than 40% of WT B cells reached division 8+ by 72 hours, IRF4-null B cells appeared to be stalled at divisions 2–5. Taken together, these data suggest that IRF4 regulates the rate of cellular proliferation during B cell differentiation that occurs around divisions 2–5 and that this role may be concentration-dependent.
Small molecule inhibitors of Activation-Induced Deaminase
Authors: Alvarez-Gonzalez Juan Antonio , Maul Robert , Kohli Rahul M. , Gearhart Patricia J. 
Published: January 2023

Abstract
Activation-Induced Deaminase (AID) is a cytosine deaminase that converts cytosine into uracil in DNA, which initiates a cascade of mutagenic DNA repair to introduce point mutations and double-strand breaks. Specific targeting of AID to the immunoglobulin heavy chain locus promotes somatic hypermutation in antibody variable genes for affinity maturation, and breaks in switch regions for class switch recombination (CSR). However, mis-targeting of AID to other loci could initiate tumor development and lead to greater drug resistance among cancer cells when continually expressed. To identify a small molecule inhibitor of AID, we screened ~400,000 compounds in conjunction with the NCATS core facility at NIH. Using FRET based analysis of cytosine deamination, we identified 150 potential inhibitors of AID catalytic activity. To confirm biological function, we examined their effects on CSR in an in vitro murine B-cell activation assay using CH12 cells, wherein 30 were confirmed inhibitor candidates. We then selected the top seven molecules to proceed with further characterization in wild type primary splenocytes, and found two near-identical compounds that had inhibitory activity. From these structures, we tested commercially available analogues and identified two molecules with inhibitory efficacy in the nanomolar range. Using these approaches, we hope to identify a small molecule with great efficacy and low toxicity for use as a molecular probe to further characterize AID’s intricacies, or even as a therapeutic agent.
IRF4 and IRF5 transcription factors exhibit shared and distinct roles in regulating human B cell differentiation and function
Authors: Shih Tiffany , De Saurav , Zhong Baohong , Barnes Betsy J. 
Published: January 2023

Abstract
Upon recognition of antigen, B cells undergo rapid proliferation followed by differentiation to specialized antibody secreting cells (ASCs). Increased levels of ASCs are seen in autoimmune diseases, such as systemic lupus erythematosus (SLE), and some B cell-associated malignancies. Studies suggest that altered transcription factor (TF) expression and/or activation play a role in the imbalance of B cell subsets in these diseases. Interferon regulatory factor 5 (IRF5) is one such TF, as polymorphisms in IRF5 associate with risk of numerous autoimmune diseases and correlate with elevated IRF5 expression in SLE patients. Recent findings from IRF5 knockdown (KD) in purified naïve B cells identified IRF5 as an early intrinsic regulator of B cell activation, proliferation and plasma cell differentiation in response to TLR9 activation. In this study, IRF4 was identified as a new IRF5 target gene by ChIP-Seq and RNA-Seq analysis. IRF4 expression is high in ASCs and murine studies show that it plays an essential, cell-intrinsic role in the generation of germinal center B cells. To date, little is known of IRF4 function in human ASC differentiation or whether these two factors (IRF4 and 5) cooperate. Activation of TLR9 and B cell receptor (BCR) revealed distinct kinetic changes in IRF4 and IRF5 during the early transition of naïve B cells to ASCs. Moreover, IRF4 KD of human primary naïve B cells resulted in significant IgD retention and reduced plasmablast (PB) differentiation without loss of early B cell activation (CD86) or key regulatory factors of class switch recombination in response to TLR9/BCR stimulation. Together, these results show that IRF4 and IRF5 are kinetically distinct TFs that have non-redundant roles in human ASC differentiation.
Immune regulation by glucocorticoids can be linked to cell-lineage-specific transcriptional responses
Authors: Franco Luis Miguel , Gadkari Manasi , Howe Katherine , Sun Jing , Kumar Parag , Kardava Lela , Biancotto Angélique , Hu Zonghui , Fraser Iain D.C. , Moir Susan , Germain Ronald N. , Tsang John S. 
Published: January 2023

Abstract
Glucocorticoids remain the most widely used class of immunosuppressive and anti-inflammatory drugs, yet significant gaps exist in our understanding of glucocorticoid-mediated immunoregulation. We have studied the transcriptional response to glucocorticoids, by total RNA sequencing and small-RNA sequencing, in nine human primary cell populations: B cells, CD4+ T cells, monocytes, neutrophils, endothelial cells, myoblasts, osteoblasts, fibroblasts, and preadipocytes. Our results suggest that the human response to glucocorticoids displays cell-lineage dependency, in terms of the individual genes and pathways that are differentially affected, as well as the magnitude and direction of transcriptional regulation. While decreased expression of genes that function in critical immune pathways is widespread, increased expression of negative regulators of immune activation is also common. Despite the critical role of B cells in many diseases for which glucocorticoids are used, glucocorticoid effects on human B cells are particularly poorly understood. We have identified significant transcriptional and functional changes in key molecules involved in B cell receptor signaling, intracellular Toll-like receptor signaling, B cell activation, differentiation, and antibody production. We have validated these results at the RNA and protein levels in vivo, by assaying B cells isolated from healthy human volunteers given a single intravenous dose of a glucocorticoid. Overall, our results illustrate the ability of an unbiased, genome-wide approach to uncover previously undescribed, cell-type-dependent transcriptional responses that can be linked to the immunoregulatory actions of glucocorticoids in humans.
Hepatic Myeloid Cell Diversity During Fibrosis and Nonalcoholic Steatohepatitis
Authors: Troutman Ty Dale , Seidman Jason S. , Link Verena M. , Hosseini Mojgan , Sakai Mashito , Bruni Cassi , Ouyang Zhengyu , Pasillas Martina , Glass Christopher K. 
Published: January 2023

Abstract
Fatty liver diseases account for rapidly growing morbidity in the United States where it is estimated that 80 to 100 million individuals have non-alcoholic fatty liver disease (NAFLD), and 6 to 16 million have the more severe variant nonalcoholic steatohepatitis (NASH). The mechanisms underlying development of NASH and progression to cirrhosis are physiologically complex. Using scRNA-Seq, we profiled CD45+ cells from the livers of mice subjected to a dietary NASH model. This approach revealed that the NASH model diet induced transcriptional diversification of Kupffer cells, and resulted in expansion of myeloid cell types. Kupffer cells, defined as F4/80High CD11bIntermediate, play a key role in the development of liver disease through their role as sentinel cells. They respond to noxious stimuli by producing chemokines and cytokines, which promote both recruitment of inflammatory leukocytes and pro-fibrogenic activation of hepatic stellate cells. Through flow cytometry, we confirmed scRNA-Seq suggestions of hepatic macrophage diversification beyond the previously reported F4/80Low CD11bHigh Ly6CHigh and F4/80Low CD11bHigh Ly6CLow recruited macrophages. Further profiling of these hepatic macrophages by RNA-Seq suggested the differential involvement of these cell subsets in the disease process. These results were corroborated using an alternative mouse model of liver fibrosis. Ongoing experiments are targeted towards assessment of the developmental origin of the distinct Kupffer cell subsets, with preliminary data suggesting a monocytic and not embryonic origin. Further efforts are focused on comparisons of myeloid populations in humans and mouse towards informing the design of future experiments.
Indispensable role of protein kinase D1 in inflammatory responses and host defense in Group B streptococci infection
Authors: Yoon Taewon , Halder Priyanka , Yi Ae-Kyung 
Published: January 2023

Abstract
Group B streptococcus (GBS) causes life-threatening diseases in neonates and immune compromised individuals. We previously found that GBS induce activation of protein kinase D1 (PKD1) in mouse and human. This PKD1 activation by GBS is dependent on the TLR signaling adaptor MyD88 and its downstream IRAK4 and IRAK1. GBS-mediated proinflammatory cytokine production is substantially suppressed by pretreatment with a PKC/PKD inhibitor Gö6976. However, it is currently unknown whether PKD1 plays a regulatory role in GBS-mediated proinflammatory responses and host defense. In the present study, to further understand contribution of PKD1 in inflammatory responses and host defense in GBS infection, tamoxifen-inducible PKD1-deficient mice were generated. We found that GBS-mediated activation of MAPKs and NF-kB and expression of cytokines/chemokines are significantly inhibited in PKD1-deficient mice. Furthermore, increased susceptibility to GBS infection was observed in PKD1-deficient mice. However, PKD1-deficient mice sensitized with D-galactosamine were more resistant to the shock-mediated death caused by antibiotic-killed GBS compared to wild-type. Our findings imply that PKD1 is one of the critical factors that plays a regulatory role in GBS-induced proinflammatory reactions and is indispensable for host defense against GBS infection and that inhibition of PKD1 activation together with antibiotic treatment in GBS-infected neonates might be an effective way to control GBS diseases.
A potential for crosstalk between inflammation, coagulation and oxidative stress host responses in malaria induced mid-gestational pregnancy loss in a rodent model
Authors: Andrew Alicer Keendalyn , Moore Julie M. , Sarr Demba , Shastri Sachi , Bracken Tara 
Published: January 2023

Abstract
Despite an overall reduction in the global malaria burden, placental malaria (PM) is a major complication of Plasmodium falciparum malaria that impacts hundreds of thousands of pregnancies annually. Primigravid women living in Sub-Saharan Africa are especially prone to PM and often experience poor maternal-fetal health outcomes though pregnancy loss, preterm delivery and infant low birth weight. Mouse models for malaria infection during pregnancy have been instrumental in our current understanding of PM, however our understanding remains incomplete. In the laboratory of Dr. Julie Moore, we have developed a mouse model of infection during pregnancy using the murine-infective species P. chaubaudi chaubaudi AS (PccAS). Infection initiated at gestation day 0 (GD 0) in mice deficient in tumor necrosis factor (TNF) function or signaling and intact C57BL/6J control mice treated with antioxidant and anticoagulant drugs reveals distinct differences in pregnancy outcome at mid-gestation (GD 10). While control mice lose their pregnancies, TNF deficient and drug-treated mice experience rescue of mid-gestational pregnancy and display improved offspring viability at sacrifice. These preliminary findings suggest that by disrupting the function of key host mediators of severe malaria, it may be possible to mitigate negative pregnancy outcomes associated with malaria. Through strategic use of TNF modified mice, combined with studies of gene expression and tissue localization, this work aims to investigate crosstalk between three major host responses to infection - inflammation, coagulation and oxidative stress – and characterize how their interplay may contribute to PM pathogenesis and associated negative pregnancy outcomes.
The host ubiquitin system in innate immunity and virus replication: proviral and antiviral functions of the host E3-ubiquitin ligase TRIM family
Authors: Rajsbaum Ricardo , Giraldo Maria Isabel , Bharaj Preeti , Atkins Colm , Xia Hongjie , Rossi Shannan L. , Lee Benhur , Shi Pei-Yong , Freiberg Alexander N. 
Published: January 2023

Abstract
Type-I Interferons (IFN-I), which are innate immune cytokines with broad-spectrum antiviral effects, are regulated by the host Ubiquitin (Ub) system. Evidence has shown that new forms of poly-Ub chains, which are not covalently attached to any protein (termed unanchored poly-Ub), are important activators of kinase activity in vitro. We recently reported that unanchored K48-linked poly-Ub synthesized by the E3-Ub ligase TRIM6 promote IFN-mediated antiviral innate immunity in vivo by activating the IKKɛ kinase. Intriguingly, we found that two different highly pathogenic viruses, Nipah (NiV; Paramyxoviridae family) and Ebola (EBOV; Filoviridae family), hijack TRIM6 to promote virus replication through mechanisms that involve both covalent and unanchored Ub. This indicates that host antiviral factors can have direct pro-viral functions. We also found that Zika virus (ZIKV; Flaviviridae family), which can cause congenital microcephaly in newborns, requires ubiquitination of viral proteins for efficient replication. Importantly, ZIKV ubiquitination occurs in vivo in a tissue-specific manner. Recombinant infectious ZIKV mutants that lack ubiquitination on specific sites are attenuated in mice and appear to differentially replicate in tissues. As compared to other flaviviruses, ZIKV acquired an additional ubiquitination site, which may be responsible for providing ZIKV an advantage to replicate in brain and reproductive tissue. This would be the first example of a flavivirus utilizing the Ub system for tissue tropism. Taken together, our work suggests that divergent viruses hijack similar innate immune host factors to enhance their replication. This information could help design novel broad-spectrum antiviral strategies
The role of Indoleamine 2,3-dioxygenase (IDO) in the survival of bone marrow resident long lived Plasma cells
Authors: Lightman Shivana M. , Carlson Louise M. , Lee Kelvin P. 
Published: January 2023

Abstract
Long lived plasma cells (LLPCs) are essential for sustained antibody responses and protective humoral immunity. How these cells maintain longevity and a durable antibody response is largely dependent on the complex nature of the bone marrow microenvironment in which these cells reside, and the pro survival factors produced in this niche. Previous work in our lab has shown that CD28 is required for LLPC survival, and we know that CD28 back signaling to CD80/86 induces IDO production from dendritic cells (DCs) within the bone marrow microenvironment. IDO is classically known to contribute to an immunosuppressive environment (specifically with respect to T cell activity). However, upon investigation of IDO in the normal setting, we observed fewer plasma cells present in the bone marrow of IDO knockout mice in comparison to wild type mice, as well as fewer antigen-specific plasma cells in the bone marrow following vaccination. Additionally, knowing the role of DCs in IDO production in the bone marrow, we employed the use of CD11c-DTR mice to observe the effects of PC survival upon DC elimination. We found that depletion of DCs significantly decreases the amount of antigen-specific plasma cells in the bone marrow. This leads us to propose a model where CD28, through back signaling to CD80/86 induces IDO production in DCs, a mechanism to maintain the bone marrow niche for long lived plasma cell survival and sustained antibody production.