bluuebunny/crossref_metadata_2025_split huggingface dataset:
A conserved salt bridge in the G-loop of multiple protein kinases is essential for catalysis, in vivo Lyn function and prevention of autoimmune disease (84.17)
Authors: Sauer Karsten , Che Jianwei , Lee Christian C. , Yang Yating , Herman Ann , Jia Yong , Velentza Anastasia , Watson James , Sternberg Luise , Kim Sunjun , Ziaee Niusha , Miller Andrew , Jackson Carie , Young Mike , Batalov Serge , Liu Yi , Warmuth Markus , Wiltshire Tim , Cooke Michael P. , Barouch-Bentov Rina 
Published: January 2023

Abstract
The glycine-rich G-loop controls ATP binding and phosphate transfer in protein kinases. Here we show that the functions of Src family and Abl protein tyrosine kinases require an electrostatic interaction between oppositely charged amino acids within their G loops that is conserved in multiple other phylogenetically distinct protein kinases from plants to humans. By limiting G-loop flexibility, it controls ATP binding, catalysis and inhibition by ATP-competitive compounds such as Imatinib. In WeeB mice, mutational disruption of the interaction results in expression of a Lyn protein with reduced catalytic activity, and in perturbed B cell receptor signaling. Like Lyn-/- mice, WeeB mice show profound defects in B cell development and function and succumb to autoimmune glomerulonephritis. This demonstrates the physiological importance of the conserved G-loop salt bridge and at the same time distinguishes the in vivo requirement for the Lyn kinase activity from other potential functions of the protein.
The tyrosine kinase c-Abl regulates CD4+ T-cell differentiation by catalyzing T-bet phosphorylation (84.5)
Authors: Lee Sang-Myeong , Chen An , Gao Beixue , Fang Deyu , Shannon Stephen , Zhang Jinping , Ye Shui Q , Calhoun Karen 
Published: January 2023

Abstract
The tyrosine kinase c-Abl is required for full activation of T cells, while its role in CD4+ T-cell differentiation has not been characterized. We report that c-Abl-deficiency skews CD4+ T cells to Th2 differentiation, and c-Abl-/- mice are more susceptible to allergic lung inflammation. c-Abl interacts with and phosphorylates T-bet, a Th1-lineage transcription factor, indicating that c-Abl is a tyrosine kinase for T-bet in T cells. c-Abl-mediated phosphorylation enhances the transcriptional activation of T-bet. Interestingly, the tyrosine residues within the T-box domain of T-bet are mapped as the predominant sites of c-Abl. Mutation of these tyrosine residues inhibits the promoter-binding activity of T-bet. Therefore, c-Abl regulates CD4+ T cell differentiation by targeting T-bet phosphorylation.
Expression of Sca-1 (Ly-6A/E) defines functionally different developmental stages of plasmacytoid dendritic cells (PDC) (84.9)
Authors: Kurig Stefanie , Fischer Jens , Dzionek Andrzej 
Published: January 2023

Abstract
Stem cell antigen-1 (Sca-1), a phosphatidylinositol-linked membrane protein is one of the most prominent stem cell markers. Functionally, Sca-1 is involved in cross-talk between lymphocyte precursors and stromal cells and plays an important role in the thymic development of T-cells and TcR-dependent activation.
Here we show that expression of Sca-1 on PDC defines functionally different developmental stages of PDC in mice. The proportion of the Sca-1+ PDC showed an organ specific variation ranging from 10-20% in BM to &gt;80% in LN. Sca-1- PDC appeared earlier in the development and give rise to Sca-1+ PDC without the need for activation. In addition to the naturally occurring up-regulation, Sca-1 expression could be further induced by TLR7 and TLR9 mediated activation. Interestingly, when exposed to CpG-oligonucleotides (TLR9), Sca-1- PDC produced significantly higher amounts of IFN-alpha. This was further confirmed by two observations: (1) after stimulation with TLR9-ligands purified PDC from BM (mainly Sca-1-) produced up to 100-fold higher amounts of IFN-alpha when compared to PDC from LN (predominantly Sca-1+), (2) Sca-1- PDC expressed significantly higher amount of genes involved in TLR mediated IFN-alpha induction, including MyD88, IRF3, IRF5, IFNAR1 and IFNAR2.
Taking together, our results identify Sca-1 as a developmental marker of PDC, which defines two functionally heterogeneous subsets of PDC.
Notch and IL-7 regulate cell fate decisions in TCRβ+ DN and ISP thymocytes (85.10)
Authors: Xiong Juan , Yankee Thomas M. 
Published: January 2023

Abstract
During the CD4-CD8- double negative (DN) stage of T cell development, the cells express TCRβ for the first time. TCRβ+ cells activate signaling pathways that promote survival, proliferation, and differentiation, but the receptors required for these signals are unknown. While Notch and IL-7 are critical for early T cell development, their function in TCRβ-expressing DN and immature single positive (ISP) CD8+ thymocytes are unclear. Here, we cultured purified DN3E (CD44-CD25hi), DN3L (CD44-CD25lo), DN4 (CD44-CD25-) and ISP CD8+ thymocytes with OP9 cells that express either the Notch ligand Delta-like 1 (DL1) or control protein. To some samples, IL-7 was also added. DN and ISP thymocytes were able to differentiate into CD4+CD8+ double positive (DP) thymocytes independently of Notch and IL-7. However, Notch promoted the proliferation and survival of the thymocytes, whereas IL-7 promoted survival and slowed differentiation. These data demonstrate that Notch and IL-7 continue to function beyond the time at which TCRβ is expressed and regulate survival, proliferation, and differentiation of TCRβ-expressing thymocytes.
This research is supported by the America Cancer Society grant RSG-08-182-01-LIB.
A full complement of CD3 ζ ITAMs is necessary for NKT cell development and function (85.11)
Authors: Becker Amy M. , Blevins Jon S. , Wight-Carter Mary , Medeiros Jennifer J. , Murray Sean , Richardson James A. , Norgard Michael V. , van Oers Nicolai S.C. 
Published: January 2023

Abstract
Invariant natural killer T (iNKT) cells express a canonical αβ TCR that recognizes glycolipids embedded in non-classical MHC molecules. These cells develop following TCR interactions with CD1d/glycolipid complexes expressed on thymocytes. The αβ TCR is non-covalently associated with six signaling proteins, CD3 ζζ and CD3 γε/δε, that signal through ITAMs. It is unknown whether a full complement of ITAMs is required for iNKT cell development and function. We previously generated CD3 ζ transgenic mice with mutations in the first (YF1,2) or all three CD3 ζ ITAMs (YF1-6) to reduce ITAM signaling. iNKT cell development was analyzed in these mice. Compared to C57BL/6 controls (0.43%; 1.04%), the percentage and absolute numbers of iNKT cells were significantly reduced in the thymus and spleen of YF1,2 (0.008%; 0.023%; 11- and 19-fold) and YF1-6 (0.044%; 0.11%, 50- and 100-fold) mice. Comparative phenotyping suggested that the iNKT cells had an early developmental block in the YF1,2 and YF1-6 mice as evidenced by an increased percentage of iNKT cells expressing CD24 compared to C57BL/6 mice. These data indicate that a full complement of CD3 ζ ITAMs is required for proper iNKT cell development. We are currently assessing the functionality of iNKT cells in the various mice in response to glycolipids or following infections with the pathogenic bacteria Borrelia burgdorferi .
The function of zbtb7b in regulating CD4 vs. CD8 T linage commitment is evolutionarily reserved (85.17)
Authors: He Xiao , Jensen Peter 
Published: January 2023

Abstract
Immature thymocytes develop into mature T lymphocytes, including 2 major functional subpopulations: CD4+ helper and CD8+ cytotoxic T cells, prior to emigration from thymus. Previously we discovered a zinc finger transcription factor, ThPOK (T helper inducing POK factor, also known as zbtb7b, a member of the zbtb7 transcription factor subfamily), functioning as a master regulator in regulating CD4 vs CD8 T cell linage commitment process. However, the molecular mechanisms underlying this process still remain to be elucidated. Intriguingly, when mouse ThPOK was ectopically expressed as a transgene in T cell lineage, all Tg mice developed fatal T lymphomas. Here we reported that when the human zbtb7b sharing an 87% identity with the mouse counterpart at DNA level and a 97% homology at protein level, was expressed in mouse T cell lineage under the CD2 promoter, some Tg mice not only could correct the ThPOK deficient phenotype (MHC class II restricted T cells are CD8+), but also developed T lymphomas. Furthermore, when another zbtb7 transcription factor subfamily member, the mouse zbtb7a sharing an 83% homology with the mouse zbtb7b at protein level, was expressed in T cells, some Tg mice again corrected the ThPOK deficient phenotype and developed T lymphomas as well. Taken together, these results indicate the function of zbtb7b in regulating CD4 vs CD8 T linage commitment is evolutionarily reserved (supported by AI-33614).
AGAPE, a new T cell specific Grb2 binding protein that plays a critical role in T cell development (85.7)
Authors: lesourne renaud , Love Paul 
Published: January 2023

Abstract
We report here the identification of a T-cell lineage protein designated AGAPE (ä-gä-pa; Another Grb2-Associated Protein Enigma). AGAPE contains a proline rich sequence that associates with the adaptor protein Grb2. It lacks any consensus catalytic domains but contains a nuclear localization sequence required for its translocation to the nucleus. AGAPE deficient mice were generated by homologous recombination in ES cells. AGAPE-/- mice have normal numbers of DN and DP thymocytes but markedly reduced numbers of CD4 SP and CD8 SP thymocytes and pheripheral T cells. Positive selection is dramatically reduced in both class II- and class I-restricted TCR transgenic AGAPE-/- mice. The proportion of CD4+CD8lo thymocytes was reduced in AGAPE-/- MHCII-/- mice suggesting that AGAPE plays and important role at the DP to SP transition stage. Interestingly, TCR induced activation responses are unaffected in AGAPE-/- thymocytes indicating that the developmental block is not due to a defect in signaling downstream of the TCR? Although DP thymocytes from AGAPE-/- mice show enhanced susceptibility to antibody-induced cell death, the introduction of the Bcl-2 transgene did not rescue T cell development. Expression of the lineage specific transcription factors ThPok and Gata-3 (but not Tox or Runx1/3) was reduced in CD69+ thymocytes purified from AGAPE-/- mice. However, enforced expression of ThPok or Gata-3 did not rescue CD4 SP development in AGAPE-/- mice. Thus, AGAPE is a new T cell specific nuclear protein that has a critical function during late thymocyte selection prior to CD4/CD8 lineage specification.
An in situ hybridization screen reveals new transcriptional regulators of third pharyngeal pouch and thymic epithelial cell differentiation. (86.12)
Authors: Condie Brian G , Wei Qiaozhi 
Published: January 2023

Abstract
Thymic epithelial cells (TECs) are the major component of the non-lymphoid framework of the thymus and are essential for normal T cell development. In mice, TECs develop from a domain of the embryonic third pharyngeal pouch endoderm marked by Foxn1 expression. Although Foxn1 is necessary for proliferation and differentiation of TECs, it is not required for specification of TECs from the pouch endoderm. Also, some aspects of TEC differentiation are Foxn1-independent. This suggests that there are additional transcription factors expressed within the third pouch endoderm that are responsible for early specification and differentiation of TECs. To identify additional transcriptional regulators of third pouch and TEC differentiation, we performed an in situ hybridization screen on a set of candidate transcription factors identified through a systematic analysis of gene expression databases. We found that most of the genes selected for detailed characterization were expressed in the third pharyngeal pouch endoderm and many of them were specifically expressed in the thymus domain of the pouch. These transcription factors are highly likely to be involved in the earliest steps of TEC development. Our work also shows that data mining of gene expression databases is an efficient way to identify genes involved in the development of a specific tissue or organ.
Estrogen receptor signaling promotes langerin+ dendritic cell differentiation by increasing expression of the transcription factor IRF4 (87.1)
Authors: Carreras-Margalef Esther , Turner Sean , Frank Mark Barton , Knowlton Nicholas , Osban Jeanette , Centola Michael B , Alberola-Ila Jose , Kovats Susan 
Published: January 2023

Abstract
During inflammation, elevated GM-CSF promotes the development of new dendritic cells (DC). We previously showed that estrogen receptor alpha (ERα) signaling in response to estradiol is required for the GM-CSF mediated differentiation of CD207/langerin+ CD11c+ tissue DC. DC development from ERα+ flt3+ committed myeloid progenitors (MP) was increased significantly after a 12 hr exposure to estradiol. We now have identified the transcription factor interferon regulatory factor 4 (IRF4) as an estradiol-induced gene in a microarray experiment comparing MP incubated -/+ estradiol. The IRF4 gene is induced by GM-CSF and critical for myeloid DC development in vivo. Quantitative PCR analysis showed that IRF4 mRNA in MP was increased two-fold within 6 hr of estradiol exposure. Differentiating CD11c- precursors also had two-fold higher levels of IRF4 protein after a 48 hr exposure to estradiol; this increase in IRF4 expression was not observed in differentiating ERα-/- CD11c- cells. Retrovirus mediated delivery of the IRF4 cDNA to ERα-/- bone marrow cells differentiating in the absence of estradiol restored the development of the estradiol-dependent DC population. Thus, ERα signaling in response to estradiol promotes the GM-CSF mediated differentiation of a langerin+ DC population by increasing expression of IRF4 protein in myeloid progenitors.
Supported by OCAST HR06-157 and NIH RR020143.
cFLIP is required for macrophage survival and neutrophil homeostasis (87.3)
Authors: Gordy Claire Lee , Pua Heather , He You-Wen 
Published: January 2023

Abstract
The anti-apoptotic protein cellular FLICE-like inhibitory protein (cFLIP) has been shown to play an important role in survival of T and B lymphocytes; however, the function of cFLIP in other immune cells is unknown. To examine the role of cFLIP in myeloid cell survival and function, we generated mice expressing a loxP-flanked cFLIP allele in the presence of Lysozyme M-Cre (cFLIPf/f Lysm-Cre). cFLIPf/f Lysm-Cre mice lack splenic marginal zone macrophages and thioglycollate-elicited peritoneal macrophages, while inflammatory monocyte and neutrophil numbers are increased. Using this model, we find that loss of cFLIP expression in macrophages and neutrophils results in a severely inflammatory phenotype, with cFLIPf/f Lysm-Cre mice displaying splenomegaly, decreased body weight, profound neutrophilia, increased production of IL-1β and GCSF, and extramedullary hematopoiesis. These defects are secondary to the loss of macrophage populations, as mixed bone marrow chimeric mice do not develop inflammation. Together, these data demonstrate a requirement for cFLIP expression for macrophage survival along with a requirement for mature macrophage populations to maintain neutrophil homeostasis in vivo.
This work is supported by an NSF Graduate Research Fellowship and NIH grants.
Crucial roles of Sp1 in the transcriptional regulation of LIGHT gene by Axl in T cell lymphoma (88.13)
Authors: Lee Eun-Hee , Park Areum , Kim Eun-Mi , Yi Ja-woon , Lee Hwa-youn , Choi Harim , Lemke Greg , Kang Hyung-Sik 
Published: January 2023

Abstract
To elucidate the role of Axl on the regulation of LIGHT expression, we established the stably over-expressing Axl T lymphoma cells in mouse EL4 (EL4-Axl) and human Jurkat T lymphoma cells (Jurkat-Axl). The expression of LIGHT and its receptor, herpes virus entry mediator (HVEM) was remarkably increased both in EL4-Axl and Jurkat-Axl compared with the individual controls. Furthermore, the expression of LIGHT, HVEM and IFN-γ was reduced in the various tissues of Axl-/- mice. The proliferation of EL4-Axl cells was inhibited by treatment of Axl-Ig in a concentration and LIGHT expression-dependent manner. In LIGHT promoter assay using a transient transfection of Jurkat-Axl cells with 5¡ deletion mutants, luciferase reporter activity showed the highest transcriptional activity in the gene segment -190/+1. Electrophoretic mobility shift assay (EMSA) and Western blot analysis revealed that Sp1 was able to directly bind to its binding site on LIGHT promoter upon triggering AKT/PI3K signaling pathway.
These results suggest that Sp1 plays a crucial role in the regulation of LIGHT expression by Axl signaling in T lymphoma.
This work was supported by the SRC/ERC program of MOST/KOSEF (RESEARCH CENTER FOR WOMEN'S DISEASES)
Increased tumorigenicity and metastasis of cervical cancer cells by HPV 16 E6-mediated blockade of lymphotoxin signaling pathway (88.14)
Authors: Kim Dong-Hern , Lee Eun-Hee , Kim Eun-Mi , Park Chang-Eon , Lee Hwa-youn , Yi Ja-woon , Yun Chul-Ho , Park Yong-Wook , Kee Seung-Jung , Kang Hyung-Sik 
Published: January 2023

Abstract
To elucidate the regulatory role of LT on human papillomavirus (HPV) E6-mediated tumorigenecity in cervical cancer, HPV 16 E6-overexpressing HeLa (HE6F) and suppressing Caski cervical cancer cells (CE6R) were established. LT gene expression was markedly suppressed in HE6F cells compared to HeLa cells, which led to a significant augmentation of HE6F cell growth via the reduction of apoptosis-associated gene expression. HPV 16 E6-induced HE6F metastasis was further amplified by silencing of LTβR using small interference RNAs. The blockade of LT signaling by LTβR-Ig treatment in HE6F cells significantly suppressed cytotoxicity mediated by NK cells. Recovery of LT by administration of LT DNA in mice challenged with TC-1 cells expressing HPV16 E6 and E7 resulted in the suppression of tumor mass, and the prolonged survival rate. Furthermore, IgA production in both fecal extracts and vaginal washes of mice administrated with LT DNA was significantly higher than that of control mice.
Taken together, these results demonstrated that progress rate of cervical cancer could be promoted by HPV E6-mediated blockade of LT signaling pathway. Regulation of LT might be an effective way to achieve stable suppression of HPV E6 oncogenesis in cervical cancer.
This work was supported by the SRC/ERC program of MOST/KOSEF (RESEARCH CENTER FOR WOMEN'S DISEASES) and by a grant from Biogreen 21 project, Rural Development Administration.
Immune-mediated depletion of pre-leukemic cells delays disease onset in a murine spontaneous leukemia model (88.17)
Authors: Reid Gregor , Seif Alix , Grupp Stephan 
Published: January 2023

Abstract
In pediatric acute leukemia, the occurrence of initiating chromosomal translocations prior to the development of leukemia provides a window of opportunity for prophylactic intervention. Successful targeting of pre-leukemic cells during the prolonged latency period may reduce or prevent the incidence of disease. We have previously reported that treatment with immunstimulatory DNA containing CpG motifs (CpG ODN) generates protective immune responses against leukemia outgrowth in a transplantable tumor model. Using the E-mu-ret transgenic mouse model of pediatric leukemia, which possesses several key features of human disease, we demonstrate a significant delay in disease onset following immune intervention in the pre-leukemic period. Specifically, the prolonged administration of CpG ODN significantly reduced the size of the pre-leukemic cell population in peripheral blood of treated mice (p = 0.028). This reduction correlated with a significant delay in disease onset in CpG ODN-treated compared to control-treated mice (p&lt;0.0001). While no mice in the control group survived beyond 250 days of age, several CpG ODN-treated mice remained disease free at one year of age and showed no sign of an expanded pre-B cell population at sacrifice. These results indicate that immune-based prevention of pediatric leukemia may be a feasible approach to reduce the impact of this disease.
Role of TRAF6 as a Regulator of Endothelial Cell Activation and Angiogenesis (88.20)
Authors: Cejas Pedro Joel , Walsh Matthew C , Pearce Erika L , Harms Gretchen , Choi Yongwon 
Published: January 2023

Abstract
Tumor angiogenesis and metastasis are dependent on tumor-secreted growth factors and pro-inflammatory signals that target receptors on endothelial cells. The signaling cascade that emanates from many of these receptors is mediated by the adaptor molecule and signal transducer TRAF6. To date, however, the intrinsic effect of TRAF6 in endothelial cell biology has not been examined in detail. Most importantly, its effect on angiogenesis, tumor growth and metastasis has not been investigated in vivo. We hypothesize that, by acting as a molecular convergence point in the signaling downstream of different pro-inflammatory and pro-angiogenic stimuli, TRAF6 is a chief regulator of endothelial cell activation and angiogenesis to drive tumor growth and metastasis.
To test this hypothesis, we have generated transgenic mice that lack TRAF6 specifically in endothelial cells (TRAF6-ΔE) to use as a novel in vivo model of angiogenesis and metastasis. Using in vitro angiogenic tests, we have found that TRAF6-deficient endothelial cells have impaired angiogenic properties, and using two different tumor models we have demonstrated that endothelial cell-expressed TRAF6 has an essential role in tumor growth.
The tyrosine kinase Syk regulates survival of chronic lymphocytic leukemia B cells through PKCδ phosphorylation and proteasome-dependent regulation of Mcl-1 expression (88.22)
Authors: Baudot Alice D , Jeandel Pierre Yves , Mouska Xavier , Maurer Ulrich , Tartare-Deckert Sophie , Raynaud Sophie D , Cassuto Jill Patrice , Ticchioni Michel , Deckert Marcel 
Published: January 2023

Abstract
B-cell chronic lymphocytic leukemia (B-CLL) is characterized by the accumulation of mature monoclonal CD5+ B cells. The disease results mainly from a failure of cells to undergo apoptosis, a process largely influenced by the existence of constitutively activated components of BCR signaling and the deregulated expression of anti-apoptotic molecules. Recent evidence pointing to a critical role of spleen tyrosine kinase (Syk) in ligand-independent BCR signaling prompted us to examine its role in CLL cells survival. Using primary leukemic cells, we show that pharmacologic inhibition of constitutive Syk activity and depletion of its expression by siRNAs led to a dramatic decrease of cell viability in all tested CLL samples (n=44/44), regardless of clinical and biological status, and induced typical apoptotic cell death, with mitochondrial failure followed by caspase 3-dependent cell death. These events were accompanied by a decreased phosphorylation of PKCδ, but not PKCα/β, and by a down-regulation of Mcl-1, but not XIAP. Moreover, the degradation of Mcl-1 following Syk inhibition relied on a caspase-independent, but proteasome-dependent process, and was rescued by the expression of a proteasome-resistant Mcl-1 mutant. Together, our results show that Syk is a key regulator of B-CLL cell viability, and represents an attractive therapeutic target.