bluuebunny/crossref_metadata_2025_split huggingface dataset:
Novel functional roles for JAML and CAR in epithelial γδ T cell-specific costimulation (135.68)
Authors: Witherden Deborah A , Verdino Petra , Rieder Stephanie E , Mills Robyn E , Teyton Luc , Fischer Wolfgang H , Wilson Ian A , Havran Wendy L 
Published: January 2023

Abstract
γδ T cells represent a major T cell population in epithelial tissues and provide a crucial first line defence against epithelial insults, yet the molecular events surrounding their activation remain poorly defined. Here we identify a γδ T cell-specific costimulatory molecule, Junctional Adhesion Molecule-Like protein (JAML). Costimulation through JAML leads to cellular proliferation, cytokine and growth factor production. Inhibition of JAML costimulation leads to diminished γδ T cell activation and delayed wound closure akin to that seen in the absence of γδ T cells. Our results identify JAML as a crucial component of epithelial γδ T cell activation. Furthermore, we show that interaction between JAML and its ligand, Coxsackie and Adenovirus receptor (CAR), extends beyond pure cell adhesion, demonstrating an important role for CAR as a signaling receptor for the immune system. This has broader implications for its proposed role in tumor migration and growth.
A critical role of IRAK1 in the differential regulation of Th17 and T regulatory cells (135.8)
Authors: Maitra Urmila , Li Liwu 
Published: January 2023

Abstract
ObjectiveIRAK-1 has been linked to several human inflammatory diseases including autoimmune diseases. Loss of IRAK1 confers resistance to EAE, however, the exact mechanism is not understood. Given the recent discovery of Th17 lineage and its link to diverse autoimmune diseases, we tested the hypothesis that IRAK1 may contribute to the progression of autoimmune diseases via modulating the differentiation of Th17 and T regulatory cells.
MethodsNaïve CD4+ T cells were isolated from splenocytes of wild type (WT) and IRAK1-/- mice. In addition to anti-CD3 and anti-CD28, CD4+ T cells were stimulated with TGF β or TGF β plus IL6. Target proteins and mRNAs were harvested and analyzed via Western blot and RT-PCR. ChIP assays were used to demonstrate in vivo binding of transcription factors to the promoter region of corresponding genes.
ResultsIn response to TGF β and IL6, loss of IRAK1 reduced IL-17A and RORγt expression in naïve CD4+ T cells contrary to WT. Also, IRAK1 knockout impaired TGFβ and IL-6 induced STAT3 Ser727 phosphorylation. ChIP assays indicated reduced in vivo STAT3 binding to the promoter of IL-17 in IRAK1-/- cells. In contrast, the levels of Foxp3 and NFAT were highly induced in IRAK1-/- cells treated with TCR agonists and TGFβ, indicating higher percentage of Treg cells. In vivo binding assays with Smad3 antibodies demonstrated increased binding to the Foxp3 promoter in IRAK1-/- cells.
ConclusionOur data supports a novel role of IRAK1 in the regulation of Th17 and Treg differentiation. These findings may have significant implications in the pathogenesis of autoimmune diseases.
This work is supported by NIH grant R01 AI64414.
Promyelocytic leukemia protein specifically augments the activation of NFAT (136.11)
Authors: Lo Yu-Hsun , Wu Chia-Chen , Shih Hsiu-Ming , Lai Ming-Zong 
Published: January 2023

Abstract
Promyelocytic leukemia (PML) protein is a tumor suppressor with complicated action mechanisms not yet fully understood. Nuclear factor of activated T cells (NFAT) is an important factor for T cell functions. In this study, we found that PML specifically enhanced the transcription activation of NFAT. In PML-null mouse embryonic fibroblasts, no transcription activity of NFAT could be detected. There was a selective requirement of PML isoform in NFAT activation: PML-I and PML-VI, but not PML-IV, increased NFAT transactivation. PML specifically promoted the expression of many, but not all, NFAT targeted genes, including TNF ƒÑ, TRAIL and Cbl-b. A specific binding of PML to NFATc was found by GST pull down assay and immuno-precipitation. The interaction of PML with NFATc in vivo was further confirmed by ChIP and DNA affinity precipitation assay (DAPA) analysis. Knockdown of PML was associated with reduced nuclear presence of NFAT. The unexpected coupling of PML with NFAT reveals a novel mechanism underlying the diverse physiological functions of PML.
Improved peripheral T cell expression of T-cell receptor (TCR)-α transgenes in TCRα/Dad1 locus-like "two gene" reporter systems (136.14)
Authors: Ortiz Benjamin D , Gomos-Klein Janette , Knirr Stefan , Andino Blanca , Harrow Faith , Erhard Karl , Kovalovsky Damian , Sant'Angelo Derek 
Published: January 2023

Abstract
The mouse T-cell receptor (TCR)-a; locus control region (LCR) resides in between the Ca; exons and the Dad1 gene. We reported the "single cell level" activity of the TCRa LCR in the context of linked heterologous single reporter transgenes randomly integrated into the genome of mice. We found that the LCR supports the normal pattern of TCRa expression in the thymus. However, abnormal ectopic expression and T cell transcript reductions were observed in the periphery. These data indicated that important cis-acting elements (not associated with the LCR) are required for strict cell type-specificity, and peripheral T cell maintenance of TCRa expression. Here we describe data from experiments with transgenic mice bearing constructs in which the LCR is flanked by two transcription units (as it is in the genome by TCRa [5'] and Dad1 [3']). In the first system, two heterologous reporter genes flank the LCR. The other system is an 80-kb TCRa/Dad1 Bacterial Artificial Chromosome (BAC) fragment in which a Va promoter driven reporter is inserted upstream of the Ca exons, followed by the TCRa LCR and the Dad1 gene locus. We find that peripheral T cells maintain expression of the TCRa reporter gene in the BAC construct. Surprisingly, in both "two-gene" reporter systems, ectopic, non-T cell expression of the TCRa reporter is virtually eliminated. The data suggest that important cis-acting elements maintaining normal TCRa expression patterns in peripheral T cells reside in the Dad 1 gene locus. Supported by NIH grants AI053050 (to BDO), RR03037 (to the Hunter College Gene Center) and UL1RR024996 (to the Clinical and Translation Science Center at Weill Cornell Medical College).
microRNA expression profiling in Multiple Sclerosis and EAE (136.21)
Authors: Escobar Sabine S , Kennedy Bryan , Kerkof Keith , Peiser Leanne , Arnett Heather A 
Published: January 2023

Abstract
microRNAs (miRNAs) are small endogenous non-coding RNAs that are emerging as key regulators of protein expression; however, little is known about the expression or roles of miRNAs in multiple sclerosis. We isolated and profiled miRNAs from normal and multiple sclerosis donors and have identified a number of miRNAs that are regulated in MS white matter and plaque tissue, as well as some which appear to have altered expression in MS grey matter tissue. Using miRNA and mRNA profiling in parallel, we have been able to identify miRNA families and predicted target networks with relevance to MS. As points of comparison to the MS tissue samples, we also extensively profiled various human immune cell subsets, as well as spinal cord samples from mice with experimental autoimmune encephalomyelitis (EAE), an animal model with similarities to MS, for miRNA expression. miRNA signatures of the whole blood human immune cell populations have enabled identification of cell-type specific miRNAs that can be used to predict the composition of the inflammatory infiltrate in the MS tissue. Identification of the regulation of miRNAs in MS tissue may lead us to a greater understanding of the pathophysiology of the disease, and potentially provide novel targets for therapeutic intervention.
Great genotypic and phenotypic diversities associated with copy-number variations (CNVs) of complement C4 and RP-C4-CYP21-TNX (RCCX) modules in Asian-Indian and European Americans (136.29)
Authors: Yu Chack-Yung , Saxena Kapil , Kitzmiller Kathryn J. , Wu Yee Ling , Zhou Bi , Esack Nazreen , Hiremath Leena , Chung Erwin K. , Yang Yan 
Published: January 2023

Abstract
Inter-individual CNVs probably afford human populations the flexibility to respond to a variety of environmental challenges, but also lead to differential disease predispositions. We investigated gene CNVs for complement C4 and steroid 21-hydroxylase from the RCCX modules located in the MHC among healthy Asian-Indian Americans (AIA) and compared them to European Americans. A combination of definitive techniques that yielded cross-confirmatory results was used. The medium gene copy-numbers for C4 and its isotypes, acidic C4A and basic C4B, were 4, 2 and 2, respectively, but their frequencies were only 53-56%. The distribution patterns for total C4 and C4A are skewed towards the high copy-number side. For example, the frequency of AIA-subjects with three copies of C4A (30.7%) was 3.92-fold of those with a single copy (7.83%). The haplotype with a single short C4B gene and the absence of C4A, which is in linkage- disequilibrium with HLA DRB1*0301 in Europeans and a strong risk factor for autoimmune diseases, has a frequency of 0.012 in AIA but 0.106 among healthy European Americans (p=6.6x10-8). The copy-number and the size of C4 genes strongly determine the plasma C4 concentrations. Parallel variations in copy-numbers of CYP21A and TNXA with total C4 were observed. The high copy-numbers of C4 and the high frequency of RCCX recombinants offer important insights to the prevalence of autoimmune and genetic diseases.
HLA genetic association analysis using the sequence feature variant type approach (136.31)
Authors: Marthandan Nishanth , Karp David R , Arnett Frank , Waller Matthew J , Guidry Paula , Feolo Michael , Marsh Steven GE , Scheuermann Richard H 
Published: January 2023

Abstract
Immunogeneticists have found significant associations between specific HLA alleles and a variety of medical conditions, including autoimmune diseases. Current disease association studies treat each HLA allele as a single complete unit, which does not illuminate the parts of the molecule associated with disease. We have developed a novel approach for genetic association analysis in which HLA genes and proteins are broken down into smaller sequence features. Sequence features are defined based on structural or functional information, and can be overlapping and continuous or discontinuous in the linear sequence. The extent of sequence variation is then assessed for each HLA sequence feature to define all variant types found in the human population. This allows for the independent analysis of disease association with any sequence feature variant type (SFVT). We tested this approach in the analysis of systemic sclerosis patients using HLA-DRB1 and HLA-DQB1 typing data. We identified a region of the HLA-DRB1 protein centered around peptide-binding pocket 7 that appears to be associated with disease risk; aromatic amino acids found in HLA-DRB1*1104 at positions 26, 28, 37 and 67 were associated with increased risk. The SFVT approach is a novel method for identifying the molecular determinants of disease association in highly polymorphic genes like the HLA.
For DAIT-DISC; supported by NIAID N01AI40076
Effector CD4+ T cells expressing IFNγ or IL-17 in pancreatic infiltrates of young NOD mice (137.23)
Authors: Li Cheng-Rui (Michael) , Bradley Linda M. 
Published: January 2023

Abstract
Type 1 diabetes (T1D) results from autoimmune destruction of insulin-producing pancreatic islet β cells. Previous studies established that the Th1 subset of CD4+ T effector/memory cells plays a central role in the induction of the aoutimmune attack against islet β cells. Using the nonobese diabetes (NOD) mouse model, our recent studies suggested that the Th17 subset CD4+ T effector cells may also play a role in T1D. In an effort to profile the kinetics of effector/memory CD4+ T cell development, we found significant percentages of CD4+ T cells that express effector/memory phenotype (CD44hi CD62L-) in peripheral lymph nodes and spleens of young female NOD mice (2 weeks old). Importantly, we were able to identify islet infiltrating CD4+ T cells in 2-week-old NOD mice, a time point when insulitis cannot be readily detected by histology. Furthermore, intracellular cytokine staining shows that significant proportions of these early infiltrating CD4+ T cells express either IFNγ or IL-17, signature cytokines of Th1 and Th17 subsets of effector/memory cells, respectively. On going efforts are being directed to elucidating how this early effector/memory population of CD4+ T cells develop and to testing the pathogenic potentials of these cells in the NOD model of T1D.
A novel protective role for the Mer receptor tyrosine kinase in glomerular injury in nephrotoxic nephritis (137.32)
Authors: Shao Wen-Hai , Rosenbaum Joshua , McGaha Tracy L. , Cohen Philip L. 
Published: January 2023

Abstract
The Mer receptor tyrosine kinase plays an important role in clearance of apoptotic cells and in modulation of innate immune responses. Mice with a Mer kinase domain deletion (Mer-KO) develop progressive lupus-like autoimmunity. Gas6, the ligand for Mer, has been detected in glomeruli. We had previously noted that normal kidneys express high levels of Mer in glomerular tissue and set out to determine its anatomical location and its function in renal tissue. We hypothesized that glomerular expression of Mer might modulate experimental immunopathology. We thus compared the susceptibility of WT and Mer-KO mice to nephrotoxic anti-glomerular basement membrane-induced murine nephrotoxic nephritis by infusing mice with 7.5 ml/kg nephrotoxic sera. Mer-KO but not WT B6 recipients showed increased proteinuria and blood urine nitrogen by day 3. Glomeruli were hyperplastic and later became necrotic. PAS-positive staining was evident in Mer-KO capillary spaces as well within tubules, consistent with massive protein leakage. This early pathological change was associated with a lower survival rate (40%) in Mer-KO mice compared to WT (100%) at day 15. Altogether, data suggest that Mer acts to preserve the kidney from immune-related inflammation, either through its role as a down regulator of cytokine production or by promoting phagocytosis of apoptotic cells. We are further investigating the role of Mer in nephrotoxic nephritis. Supported by NIDCR and the US Department of Veterans Affairs.
The Dot1L histone H3 methyltransferase is essential for early Mammalian development and hematopoiesis (138.15)
Authors: Fields Patrick E. , Feng Yi , Yang Yanping , Ortega Manoela M. , Copeland Jessica , Vivian Jay L. 
Published: January 2023

Abstract
Histone methylation of H3 is catalyzed by at least two families of proteins, the PRMT family and the SET family. Recent studies have found a new histone H3 methyltransferase without a SET domain named Dot1L (disrupter of telomere silencing 1-like). To investigate the function of Dot1L, a Dot1L deficient mouse line was generated by injection of murine embryonic stem (ES) cells containing a gene trap into C57BL/6 female blastocysts (clone #RRR032; Bay Genomics, San Francisco, CA). Fibroblasts from Dot1L-homozygous mutant embryos were completely defective in their ability to methylate lysine 79 of histone H3. All knockout embryos died by embryonic day 13. The knockout embryos showed significant reductions in blood and blood vessel formation. Further, colony assays revealed a defect primarily in early erythropoiesis. The early erythroid precursor cells were defective in their ability to respond to erythropoietin, and the cells were more susceptible to early apoptosis after stimulation. We conclude that Dot1L plays an important role in early hematopoiesis, primarily affecting early erythroid precursor cells.
MyD88 expression in stem cells regulates hematopoietic reconstitution following bone marrow transplantation (138.16)
Authors: Rahman Adeeb H , Turka Laurence A 
Published: January 2023

Abstract
Myeloid differentiation protein 88 (MyD88) is an adaptor protein that is required for signaling through most Toll-like receptors (TLRs) as well as the IL-1R family. While these pathways are primarily considered in cells of the innate immune system, TLRs are also expressed by hematopoietic stem cells (HSCs) and progenitor cells. We sought to examine the potential importance of MyD88 during hematopoietic development. We find that bone marrow from WT and Myd88-/- C57BL/6 mice contains comparable numbers of HSCs, which give rise to differentiated progenitors with equivalent efficiency in vitro. However, following a competitive mixed bone marrow transplant (BMT) into lethally irradiated recipients, WT HSCs display a significant advantage over Myd88-/- HSCs, resulting in a stable predominance of WT-derived cells in the reconstituted myeloid and lymphoid compartments. This skewed ratio of mature WT to Myd88-/- cells in the periphery of reconstituted mice is apparent in all developmental stages including the earliest long term HSCs. Interestingly, we find that WT HSCs do not exhibit this advantage over Myd88-/- HSCs following competitive BMT into non-irradiated recipients. Overall, our data suggest that factors associated with irradiation, such as inflammation, act on HSCs in a MyD88-dependent fashion to promote long-term engraftment of transplanted bone marrow.
Nuclear factor TOX is required for lymph node organogenesis and NK cell development (138.29)
Authors: Aliahmad Parinaz , Kaye Jonathan 
Published: January 2023

Abstract
Thymocyte selection-associated HMG-box protein (TOX) is a nuclear DNA-binding factor that is expressed at discrete stages of T cell development in the thymus. We have previously reported that development of CD4 T cells, NKT cells and T regulatory cells is severely inhibited in TOX-deficient mice, as a result of a block in the completion of thymic positive selection. In contrast, CD8 T cells, γδT cells, and B cells develop in these animals. Here we document that there are other severe abnormalities in development of the immune system in the absence of TOX, including inhibition of the development of NK cells and loss of lymph nodes (LN). TOX-deficient bone marrow precursor cells fail to develop into NK cells under appropriate conditions in vitro, suggesting an intrinsic defect in hematopoiesis. Peripheral lymph nodes are undetectable in TOX-deficient mice, while the number and size of Peyer's patches are severely reduced. Bone marrow chimeras demonstrate that loss of LNs is not due to aberrant migration of lymphoid populations. Indeed, TOX-deficient mice have few phenotypically defined lymphoid tissue inducer cells, suggesting a developmental defect in organogenesis. We are currently investigating the mechanisms surrounding these phenotypes.
c-Myb is crucial for B-lymphocyte development (138.30)
Authors: Bender Timothy P , Allman David M , Rajewsky Klaus , Fahl Shawn P 
Published: January 2023

Abstract
c-Myb is abundantly expressed in lymphocyte progenitors yet little is known about c-Myb function during lymphocyte development due to the embryonic lethality of Myb null mutations. We have targeted the murine Myb locus with loxP sites for conditional deletion and crossed Mybff mice to the Mb1Cre and CD19Cre strains to ablate c-Myb expression during different stages of B cell development. Mb1Cre mediated deletion begins in pre-pro-B cells and is complete by the pro-B cell stage. We detect a no decrease in the number pre-pro-B cells but &lt;1% the normal number of CD19+ B-lineage cells at any subsequent stage of development, demonstrating that c-Myb is essential for the development past the point of commitment to the B cell lineage. In Mybf/f CD19Cre mice, B-lineage cells are reduced in number by 80% and blocked in transition from pro-B to pre-B cells, resulting in about 10% the normal number of IgM+ bone marrow B cells. c-Myb deficient B cells are hyporesponsive to IL-7 in methylcellulose cloning assays and in vitro proliferation assays, suggesting that at least part of the defect in pro-B to pre-B cell transition is due to a defect in the IL-7 signaling pathway.
IgE Receptor Activates Stat5 via a Fyn-kinase Pathway (139.10)
Authors: Barnstein Brian , Ryan John 
Published: January 2023

Abstract
We have recently found that the transcription factor Stat5 is rapidly activated by IgE crosslinkage, and that its expression is critical to IgE-mediated degranulation, LTB4 production, cytokine secretion, and survival signals. Stat5KO mast cells induced cytokine mRNAs normally following IgE crosslinkage, but these mRNAs were not sustained over time, and were degraded at twice the rate observed in wild type cells. We find that Stat5 is associated with the IgE receptor after antigen-mediated activation. Moreover, cholesterol depletion or Fyn kinase deletion prevents Stat5 tyrosine phosphorylation in response to IgE crosslinkage. In contrast, the Lyn, Syk, MAPK, and PI-3 kinase pathways do not appear to regulate Stat5 tyrosine phosphorylation via the IgE receptor. We also find that Stat5 is serine phosphorylated during IgE signaling. These data suggest that Stat5 is activated through a lipid raft- and Fyn-dependent pathway. Our work demonstrates that Stat5 is a critical factor in IgE-induced mast cell activation that could provide a new clinical target for mast cell-associated inflammatory diseases. Supported by NIH grants 1R01 AI59638 and U19A1077435.
High Resolution Imaging Methods to Probe FcεRI Membrane Organization & Behavior (139.6)
Authors: Wilson Bridget S , Lidke Diane , Lidke Keith , Andrews Nicholas , Murphy Robert , Oliver Janet M 
Published: January 2023

Abstract
We have applied a combination of high resolution microscopy approaches, together with lipid mass spec analysis, to study the membrane organization of high affinity IgE receptors (FcεRI), their signaling partners and the local lipid environment. Using immunogold labeling and transmission electron microscopy (TEM) imaging of fixed membrane sheets, we find resting receptors in singlets and small clusters. After crosslinking, clusters increase dramatically in size and recruit specific downstream signaling molecules to form primary signaling domains. Atomic Force Microscopy (AFM) studies provide maps of topographical features of the cytoplasmic face of the membrane and demonstrate the importance of cholesterol to the height of these features. Consistent with this, lipid mass spec analysis of isolated FcεRI membrane patches show that 50% of the lipid surrounding receptors is cholesterol. Recently, we have generated novel quantum dot (QD) based probes to study the dynamics of early events in FcεRI signaling. Monovalent IgE-QD complexes are used to track the lateral motion of individual FcεRI in resting and activated states. These studies document transient co-confinement of resting IgE-bound receptors, as well as the immobilization of receptors after extensive crosslinking. Mathematical models show that clusters form when receptors rapidly exchange between domains or confinement zones, consistent with experimental results. The motion of QD-IgE has also been tracked in live cells transfected with GFP-actin using total internal reflection microscopy (TIRF), providing direct evidence that actin corrals provide one mechanism for defining confinement zones. Supported by NIH RO1 AI051575 (BW), NIH RO1 GM49814 (JM0) and Sandia Labs (LDRD).