Transcription Factor STAT3 Research Articles

A STAT5-Smad3 dyad regulates adipogenic plasticity of visceral adipose mesenchymal stromal cells during chronic inflammation

Adipogenic differentiation of visceral adipose tissue-resident multipotent mesenchymal stromal cells (VA-MSC) into adipocytes is metabolically protective. Under chronic inflammatory stress, this neoadipogenesis process is suppressed by various pro-inflammatory cytokines and growth factors. However, the underlying mechanism(s) regulating VA-MSC plasticity remains largely unexplored. Using an adipogenic differentiation screen, we identified IFNγ and TGFβ as key inhibitors of primary human VA-MSC differentiation. Further studies using human and mouse VA-MSCs and a chronic high-fat diet-fed murine model revealed that IFNγ/JAK2-activated STAT5 transcription factor is a central regulator of VA-MSC differentiation under chronic inflammatory conditions. Furthermore, our results indicate that under such conditions, IFNγ-activated STAT5 and TGFβ-activated Smad3 physically interact via Smad4. This STAT5–Smad4-Smad3 complex plays a crucial role in preventing the early adipogenic commitment of VA-MSCs by suppressing key pro-adipogenic transcription factors, including CEBPδ, CEBPα, and PPARγ. Genetic or pharmacological disruption of IFNγ-TGFβ synergy by inhibiting either STAT5 or Smad3 rescued adipogenesis under chronic inflammatory stress. Overall, our study delineates a central mechanism of MSC plasticity regulation by the convergence of multiple inflammatory signaling pathways.

Hypoxia-Induced Nestin Regulates Viability and Metabolism of Lung Cancer by Targeting Transcriptional Factor Nrf2, STAT3, and SOX2.

Objective To investigate hypoxia-induced Nestin regulates lung cancer viability and metabolism by targeting transcription factors Nrf2, STAT3, and SOX2. Methods Eighty-four cases of nonsmall cell lung cancer (nonsmall cell lung cancer, NSCLC), which had been treated from June 2020 to February 2021, were randomly selected from our clinicopathology database. Immunohistochemical staining of collected tissue cells was performed to assess the expression patterns of Nestin, STAT3, Nrf2, and SOX2. Data were quantified and statistically analyzed using one-way and two-way ANOVA tests with P < 0.05. Results Clinicopathological findings showed significant differences in lymph node metastasis, tissue differentiation, and histology on induction of Nestin expression; Nestin expression correlated with STAT3, Nrf2, and SOX2 expression.Nestin/STAT3/SOX2/Nrf2 are involved in angiogenesis and lung cancer development. Conclusion Hypoxia-induced Nestin promotes the progression of nonsmall lung cancer cells by targeting the downstream transcription factors STAT3, Nrf2, and SOX2.

Engineered endosymbionts that alter mammalian cell surface marker, cytokine and chemokine expression

Developing modular tools that direct mammalian cell function and activity through controlled delivery of essential regulators would improve methods of guiding tissue regeneration, enhancing cellular-based therapeutics and modulating immune responses. To address this challenge, Bacillus subtilis was developed as a chassis organism for engineered endosymbionts (EES) that escape phagosome destruction, reside in the cytoplasm of mammalian cells, and secrete proteins that are transported to the nucleus to impact host cell response and function. Two synthetic operons encoding either the mammalian transcription factors Stat-1 and Klf6 or Klf4 and Gata-3 were recombined into the genome of B. subtilis expressing listeriolysin O (LLO) from Listeria monocytogenes and expressed from regulated promoters. Controlled expression of the mammalian proteins from B. subtilis LLO in the cytoplasm of J774A.1 macrophage/monocyte cells altered surface marker, cytokine and chemokine expression. Modulation of host cell fates displayed some expected patterns towards anti- or pro-inflammatory phenotypes by each of the distinct transcription factor pairs with further demonstration of complex regulation caused by a combination of the EES interaction and transcription factors. Expressing mammalian transcription factors from engineered intracellular B. subtilis as engineered endosymbionts comprises a new tool for directing host cell gene expression for therapeutic and research purposes.

STAT1 Contributes to Microglial/Macrophage Inflammation and Neurological Dysfunction in a Mouse Model of Traumatic Brain Injury.

Traumatic brain injury (TBI) triggers a plethora of inflammatory events in the brain that aggravate secondary injury and impede tissue repair. Resident microglia (Mi) and blood-borne infiltrating macrophages (MΦ) are major players of inflammatory responses in the post-TBI brain and possess high functional heterogeneity. However, the plasticity of these cells has yet to be exploited to develop therapies that can mitigate brain inflammation and improve the outcome after TBI. This study investigated the transcription factor STAT1 as a key determinant of proinflammatory Mi/MΦ responses and aimed to develop STAT1 as a novel therapeutic target for TBI using a controlled cortical impact model of TBI on adult male mice. TBI induced robust upregulation of STAT1 in the brain at the subacute injury stage, which occurred primarily in Mi/MΦ. Intraperitoneal administration of fludarabine, a selective STAT1 inhibitor, markedly alleviated proinflammatory Mi/MΦ responses and brain inflammation burden after TBI. Such phenotype-modulating effects of fludarabine on post-TBI Mi/MΦ were reproduced by tamoxifen-induced, selective KO of STAT1 in Mi/MΦ (STAT1 mKO). By propelling Mi/MΦ away from a detrimental proinflammatory phenotype, STAT1 mKO was sufficient to reduce long-term neurologic deficits and brain lesion size after TBI. Importantly, short-term fludarabine treatment after TBI elicited long-lasting improvement of TBI outcomes, but this effect was lost on STAT1 mKO mice. Together, our study provided the first line of evidence that STAT1 causatively determines the proinflammatory phenotype of brain Mi/MΦ after TBI. We also showed promising preclinical data supporting the use of fludarabine as a novel immunomodulating therapy to TBI.SIGNIFICANCE STATEMENT The functional phenotype of microglia and macrophages (Mi/MΦ) critically influences brain inflammation and the outcome after traumatic brain injury (TBI); however, no therapies have been developed to modulate Mi/MΦ functions to treat TBI. Here we report, for the first time, that the transcription factor STAT1 is a key mediator of proinflammatory Mi/MΦ responses in the post-TBI brain, the specific deletion of which ameliorates neuroinflammation and improves long-term functional recovery after TBI. We also show excellent efficacy of a selective STAT1 inhibitor fludarabine against TBI-induced functional deficits and brain injury using a mouse model, presenting STAT1 as a promising therapeutic target for TBI.

Dissecting the dynamic transcriptional landscape of early T helper cell differentiation into Th1, Th2, and Th1/2 hybrid cells.

Selective differentiation of CD4+ T helper (Th) cells into specialized subsets such as Th1 and Th2 cells is a key element of the adaptive immune system driving appropriate immune responses. Besides those canonical Th-cell lineages, hybrid phenotypes such as Th1/2 cells arise in vivo, and their generation could be reproduced in vitro. While master-regulator transcription factors like T-bet for Th1 and GATA-3 for Th2 cells drive and maintain differentiation into the canonical lineages, the transcriptional architecture of hybrid phenotypes is less well understood. In particular, it has remained unclear whether a hybrid phenotype implies a mixture of the effects of several canonical lineages for each gene, or rather a bimodal behavior across genes. Th-cell differentiation is a dynamic process in which the regulatory factors are modulated over time, but longitudinal studies of Th-cell differentiation are sparse. Here, we present a dynamic transcriptome analysis following Th-cell differentiation into Th1, Th2, and Th1/2 hybrid cells at 3-h time intervals in the first hours after stimulation. We identified an early bifurcation point in gene expression programs, and we found that only a minority of ~20% of Th cell-specific genes showed mixed effects from both Th1 and Th2 cells on Th1/2 hybrid cells. While most genes followed either Th1- or Th2-cell gene expression, another fraction of ~20% of genes followed a Th1 and Th2 cell-independent transcriptional program associated with the transcription factors STAT1 and STAT4. Overall, our results emphasize the key role of high-resolution longitudinal data for the characterization of cellular phenotypes.

MiR-150 in HTLV-1 infection and T-cell transformation.

Human T-cell leukemia virus-1 (HTLV-1) is a retrovirus that persistently infects CD4+ T-cells, and is the causative agent of adult T-cell leukemia/lymphoma (ATLL), tropical spastic paraparesis/HTLV-1-associated myelopathy (TSP/HAM) and several inflammatory diseases. T-cell transformation by HTLV-1 is driven by multiple interactions between viral regulatory proteins and host cell pathways that govern cell proliferation and survival. Studies performed over the last decade have revealed alterations in the expression of many microRNAs in HTLV-1-infected cells and ATLL cells, and have identified several microRNA targets with roles in the viral life cycle and host cell turnover. This review centers on miR-150-5p, a microRNA whose expression is temporally regulated during lymphocyte development and altered in several hematological malignancies. The levels of miR-150-5p are reduced in many HTLV-1-transformed- and ATLL-derived cell lines. Experiments in these cell lines showed that downregulation of miR-150-5p results in activation of the transcription factor STAT1, which is a direct target of the miRNA. However, data on miR-150-5p levels in freshly isolated ATLL samples are suggestive of its upregulation compared to controls. These apparently puzzling findings highlight the need for more in-depth studies of the role of miR-150-5p in HTLV-1 infection and pathogenesis based on knowledge of miR-150-5p-target mRNA interactions and mechanisms regulating its function in normal leukocytes and hematologic neoplasms.

Deciphering the dynamic niches and regeneration-associated transcriptional program of motoneurons following peripheral nerve injury

SummaryRobust axon regeneration of motoneurons (MNs) occurs in rodent models upon peripheral nerve injury (PNI). However, genome-wide dynamic molecules and permissive microenvironment following insult in MNs remain largely unknown. Here, we firstly tackled by high-coverage and massive sequencing of laser-dissected individual ChAT+ cells to uncover molecules and pro-regenerative programs of MNs from injury to the regenerating phase after PNI. “Injured” populations at 1d∼7d were well distinguished and three response phases were well defined by elucidating with several clues (Gap43, etc). We found remarkable changes of genes expressed by injured motoneurons to activate and enhance intrinsic axon regrowth or crosstalk with other cellular or non-cellular counterpart in the activated regenerative microenvironment, specifically microglia/macrophage. We also identified an injury and regeneration-associated module and critical regulators including core transcription factors (Atf3, Arid5a, Klf6, Klf7, Jun, Stat3, and Myc). This study provides a vital resource and critical molecules for studying neural repair of axotomized motoneurons.

STAT3-mediated upregulation of LINC00520 contributed to temozolomide chemoresistance in glioblastoma by interacting with RNA-binding protein LIN28B

A considerable number of glioblastoma (GBM) patients developed drug resistance to Temozolomide (TMZ) during chemotherapy, resulting in therapeutic failure and tumor recurrence. However, the exact mechanism of TMZ chemoresistance in GBM is still poorly clarified. As a novel identified lncRNA, LINC00520 was located on chromosome 14 and overexpressed in multiple human cancers. This study was designed and conducted to investigate the role and underlying mechanism of LINC00520 in GBM chemoresistance to TMZ. The qRT-PCR assay demonstrated that LINC00520 was significantly overexpressed in TMZ-sensitive and/or TMZ-resistant GBM cells (P < 0.001). The silencing of LINC00520 markedly reduced the cell viability, suppressed colony formation, induced cell apoptosis and G1/S phase arrest in TMZ-resistant cells (P < 0.001). In contrast, overexpression of LINC00520 conferred TMZ-resistant phenotype of GBM cells in vitro (P < 0.001). The orthotopic xenograft model was established and the results indicated that the volume of tumor xenografts in vivo was markedly inhibited by TMZ treatment after the silencing of LINC00520 (P < 0.001). Luciferase reporter assay and chromatin immunoprecipitation (ChIP) assay revealed a strong affinity of transcription factor STAT3 to the promoter regions of LINC00520, suggesting that STAT3 mediated the aberrant expression of LINC00520 in GBM. Further experiments demonstrated that LINC00520 could interact with RNA-binding protein LIN28B to inhibit autophagy and reduce DNA damage, thereby contributing to TMZ chemoresistance in GBM. These findings suggested that STAT3/LINC00520/LIN28B axis might be a promising target to improve TMZ chemoresistance of GBM.

Free-cell therapeutics and mechanism of exosomes from adipose-derived stem cells in promoting wound healing: current understanding and future applications.

Free-cell therapeutics and mechanism of exosomes from adipose-derived stem cells in promoting wound healing: current understanding and future applications.

Front Cover: Resveratrol Attenuates Mast Cell Mediated Allergic Reactions: Potential for Use as a Nutraceutical in Allergic Diseases?

Mol. Nutr. Food Res. 2022, 66, 202200170 DOI: 10.1002/mnfr.202200170 Resveratrol, a polyphenol found in red grapes, inhibits release of pre-stored and newly synthesized pro-inflammatory mediators in mast cells activated by various stimuli. Signaling pathways inhibited by resveratrol include among others MAP kinases ERK and p38 as well as transcription factors STAT3 and NF-κB. This is reported by Axel Lorentz and co-workers in article number 2200170.

Impaired type I interferon signaling activity implicated in the peripheral blood transcriptome of preclinical Alzheimer's disease.

SummaryBackgroundSubjective or objective subtle cognitive decline (SCD) is considered the preclinical manifestation of Alzheimer's disease (AD), which is a potentially crucial window for preventing or delaying the progression of the disease.MethodsTo explore the potential mechanism of disease progression and identify relevant biomarkers, we comprehensively assessed the peripheral blood transcriptomic alterations in SCD, covering lncRNA, mRNA, and miRNA.FindingsDysregulated protein-coding mRNA at both gene and isoform levels implicated impairment in the type I interferon signaling pathway in SCD. Specifically, this pathway was regulated by the transcription factor STAT1 and ncRNAs NRIR and has-miR-146a-5p. The miRNA-mRNA-lncRNA co-expression network revealed hub genes for the interferon module. Individuals with lower interferon signaling activity and lower expression of a hub gene STAT1 exhibited a higher conversion rate to mild cognitive impairment (MCI).InterpretationOur findings illustrated the down-regulation of interferon signaling activity would potentially increase the risk of disease progression and thus serve as a pre-disease biomarker.FundingThis work was partly supported by National Key R&D Program of China (2020YFA0712403), National Natural Science Foundation of China (61932008), Shanghai Municipal Science and Technology Major Project (2018SHZDZX01), the 111 Project (No. B18015) of China, Greater Bay Area Institute of Precision Medicine (Guangzhou) (Grand No. IPM21C008), Natural Science Foundation of Shanghai (21ZR1403200), and Shanghai Center for Brain Science and Brain-Inspired Technology.

Plasmalogen-Mediated Activation of GPCR21 Regulates Cytolytic Activity of NK Cells against the Target Cells.

It is widely known that the immune system becomes slower to respond among elderly people, making them more susceptible to viral infection and cancer. The mechanism of aging-related immune deficiency remained mostly elusive. In this article, we report that plasmalogens (Pls), special phospholipids found to be reduced among the elderly population, critically control cytolytic activity of human NK cells, which is associated with activation of a cell surface receptor, G protein-coupled receptor 21 (GPCR21). We found the extracellular glycosylation site of GPCR21, which is conserved among the mammalian species, to be critically important for the activation of NK cells by Pls. The Pls-GPCR21 signaling cascade induces the expression of Perforin-1, a cytolytic pore-forming protein, via activation of STAT5 transcription factor. Inhibition of STAT5 abrogates GPCR21-mediated cytolytic activation of NK cells against the target cancer cells. In addition, oral ingestion of Pls inhibited cancer growth in SCID mice and inhibited the systemic spread of murine CMV in adult C57BL/6J mice. These findings advocate that Pls-GPCR21 signaling could be critical in maintaining NK cell function, and that the age-related reduction of this signaling cascade could be one of the factors behind immune deficiency in mammals, including humans.

Paeonol protects against doxorubicin-induced cardiotoxicity by promoting Mfn2-mediated mitochondrial fusion through activating the PKCε-Stat3 pathway

IntroductionThe anti-cancer medication doxorubicin (Dox) is largely restricted in clinical usage due to its significant cardiotoxicity. The only medication approved by the FDA for Dox-induced cardiotoxicity is dexrazoxane, while it may reduce the sensitivity of cancer cells to chemotherapy and is restricted for use. There is an urgent need for the development of safe and effective medicines to alleviate Dox-induced cardiotoxicity. ObjectivesThe objective of this study was to determine whether Paeonol (Pae) has the ability to protect against Dox-induced cardiotoxicity and if so, what are the underlying mechanisms involved. MethodsSprague-Dawley rats and primary cardiomyocytes were used to create Dox-induced cardiotoxicity models. Pae's effects on myocardial damage, mitochondrial function, mitochondrial dynamics and signaling pathways were studied using a range of experimental methods. ResultsPae enhanced Mfn2-mediated mitochondrial fusion, restored mitochondrial function and cardiac performance both in vivo and in vitro under the Dox conditions. The protective properties of Pae were blunted when Mfn2 was knocked down or knocked out in Dox-induced cardiomyocytes and hearts respectively. Mechanistically, Pae promoted Mfn2-mediated mitochondria fusion by activating the transcription factor Stat3, which bound to the Mfn2 promoter in a direct manner and up-regulated its transcriptional expression. Furthermore, molecular docking, surface plasmon resonance and co-immunoprecipitation studies showed that Pae’s direct target was PKCε, which interacted with Stat3 and enabled its phosphorylation and activation. Pae-induced Stat3 phosphorylation and Mfn2-mediated mitochondrial fusion were inhibited when PKCε was knocked down. Furthermore, Pae did not interfere with Dox's antitumor efficacy in several tumor cells. ConclusionPae protects the heart against Dox-induced damage by stimulating mitochondrial fusion via the PKCε-Stat3-Mfn2 pathway, indicating that Pae might be a promising therapeutic therapy for Dox-induced cardiotoxicity while maintaining Dox's anticancer activity.

A Novel Mechanism of Coactivator Recruitment by the Nurr1 Nuclear Receptor

Nuclear receptors constitute one of the largest families of transcription factors that regulate genes in metazoans in response to small molecule ligands. Many receptors harbor two transactivation domains, one at each end of the protein sequence. Whereas the molecular mechanisms of transactivation mediated by the ligand-binding domain at the C-terminus of the protein are generally well established, the mechanism involving the N-terminal domain called activation function 1 (AF1) has remained elusive. Previous studies implicated the AF1 domain as a significant contributor towards the overall transcriptional activity of the NR4A family of nuclear receptors and suggested that the steroid receptor coactivators (SRCs) play an important role in this process. Here we show that a short segment within the AF1 domain of the NR4A receptor Nurr1 can directly engage with the SRC1 PAS-B domain. We also show that this segment forms a helix upon binding to a largely hydrophobic groove on PAS-B, overlapping with the surface engaged by the STAT6 transcription factor, suggesting that this mode of recruitment could be shared by diverse transcription factors including other nuclear receptors.

Anti-Inflammatory Properties of Metformin During Cultivation of Primary Rat Astrocytes in a Medium with High Glucose Concentration.

Investigation of the relationship between inflammation and energy metabolism is important for understanding biology of chronic noncommunicable diseases. Use of metformin, a drug for treatment of diabetes, is considered as a promising direction for treatment of neurodegenerative diseases and other neuropathologies with an inflammatory component. Astrocytes play an important role in the regulation of energy metabolism and neuroinflammation; therefore, we studied the effect of metformin on the cellular responses of primary rat astrocytes cultured in a medium with high glucose concentration (22.5 mM, 48-h incubation). Lipopolysaccharide (LPS) was used to stimulate inflammation. The effects of metformin were assessed by monitoring changes in the expression of proinflammatory cytokines and synthesis of oxylipins, assayed with ultra-high-performance liquid chromatography and tandem mass spectrometry (UPLC-MS/MS). Changes at the intracellular level were assessed by analyzing phosphorylation of ERK kinase and transcription factor STAT3, as well as enzymes mediating oxylipin synthesis, cyclooxygenase 1 and 2 (COX). It was found that, independent on glucose concentration, metformin reduced the LPS-stimulated release of cytokines IL-1β and IL-6, decreased activity of the transcription factor STAT3, ERK kinase, synthesis of the derivatives of the cyclooxygenase branch of metabolism of oxylipins and anandamide, and did not affect formation of ROS. The study of energy phenotype of the cells showed that metformin activated glycolysis and inhibited mitochondrial respiration and oxidative phosphorylation, independent on LPS stimulation and cell cultivation at high glucose concentration. Thus, it has been shown that metformin exhibits anti-inflammatory effects, and its effect on the synthesis of cytokines, prostaglandins, and other lipid mediators could determine beneficial effects of metformin in models of neuropathology.

Selective Pharmaceutical Inhibition of PARP14 Mitigates Allergen-Induced IgE and Mucus Overproduction in a Mouse Model of Pulmonary Allergic Response.

The type 2 cytokines IL-4 and IL-13, which share use of an IL-4 receptor α-chain and its nuclear induction of the transcription factor STAT6, are crucial in elicitation and maintenance of allergic conditions including asthma. STAT6 binds poly(ADP-ribose) polymerase (PARP)14, an ADP-ribosyl monotransferase. Elimination of PARP14 by gene targeting led to attenuation of OVA-specific allergic lung inflammation. However, PARP14 has multiple functional domains apart from the portion that catalyzes ADP-ribosylation, and it is not clear whether inhibition of the catalytic function has any biological consequence. Using BALB/c mice sensitized to the allergen Alternaria alternata, we show that peroral administration of RBN012759, a highly selective inhibitor of ADP-ribosylation by PARP14 with negligible impact on other members of the PARP gene family, achieved biologically active plasma concentrations and altered several responses to the Ag. Specifically, the pharmaceutical compound decreased mucus after allergen challenge, blunted the induced increases in circulating IgE, and prevented suppression of IgG2a. We conclude that PARP14 catalytic activity can contribute to pathogenesis in allergic or atopic processes and propose that other biological endpoints dependent on ADP-ribosylation by PARP14 can be targeted using selective inhibition.

Frequency distribution of cytokine and associated transcription factor single nucleotide polymorphisms in Zimbabweans: Impact on schistosome infection and cytokine levels.

Cytokines mediate T-helper (TH) responses that are crucial for determining the course of infection and disease. The expression of cytokines is regulated by transcription factors (TFs). Here we present the frequencies of single nucleotide polymorphisms (SNPs) in cytokine and TF genes in a Zimbabwean population, and further relate SNPs to susceptibility to schistosomiasis and cytokine levels. Individuals (N = 850) were genotyped for SNPs across the cytokines IL4, IL10, IL13, IL33, and IFNG, and their TFs STAT4, STAT5A/B, STAT6, GATA3, FOXP3, and TBX21 to determine allele frequencies. Circulatory levels of systemic and parasite-specific IL-4, IL-5, IL-10, IL-13, and IFNγ were quantified via enzyme-linked immunosorbent assay. Schistosoma haematobium infection was determined by enumerating parasite eggs excreted in urine by microscopy. SNP allele frequencies were related to infection status by case-control analysis and logistic regression, and egg burdens and systemic and parasite-specific cytokine levels by analysis of variance and linear regression. Novel findings were i) IL4 rs2070874*T’s association with protection from schistosomiasis, as carriage of ≥1 allele gave an odds ratio of infection of 0.597 (95% CIs, 0.421–0.848, p = 0.0021) and IFNG rs2069727*G’s association with susceptibility to schistosomiasis as carriage of ≥1 allele gave an odds ratio of infection of 1.692 (1.229–2.33, p = 0.0013). Neither IL4 rs2070874*T nor IFNG rs2069727*G were significantly associated with cytokine levels. This study found TH2-upregulating SNPs were more frequent among the Zimbabwean sample compared to African and European populations, highlighting the value of immunogenetic studies of African populations in the context of infectious diseases and other conditions, including allergic and atopic disease. In addition, the identification of novel infection-associated alleles in both TH1- and TH2-associated genes highlights the role of both in regulating and controlling responses to Schistosoma.

Evaluation of the putative lymphoma-associated point mutation D427H in the STAT3 transcription factor

BackgroundSignal transducer and activator of transcription 3 (STAT3) is an oncogenic transcription factor that promotes cell proliferation and immunomodulation in untransformed cells and maintains stemness of transformed cells, facilitating invasion and metastasis. Numerous point mutations in the STAT3 protein have been identified that drive malignancy in various tumor entities. The missense mutation D427H localized in the STAT3 DNA-binding domain has been previously reported in patients with NK/T cell lymphomas. To assess the biological activity of this missense mutation, we compared the STAT3-D427H mutant to wild-type (WT) protein as well as the known hyper-active mutant F174A.ResultsAlthough previously reported as an activating mutation, the STAT3-D427H mutant neither showed elevated cytokine-induced tyrosine phosphorylation nor altered nuclear accumulation, as compared to the WT protein. However, the D427H mutant displayed enhanced binding to STAT-specific DNA-binding sites but a reduced sequence specificity and dissociation rate from DNA, which was demonstrated by electrophoretic mobility shift assays. This observation is consistent with the phenotype of the homologous E421K mutation in the STAT1 protein, which also displayed enhanced binding to DNA but lacked a corresponding increase in transcriptional activity.ConclusionsBased on our data, it is unlikely that the D427H missense mutation in the STAT3 protein possesses an oncogenic potential beyond the WT molecule.

STAT1 signaling protects self-reactive T cells from control by innate cells during neuroinflammation.

The transcription factor STAT1 plays a critical role in modulating the differentiation of CD4+ T cells producing IL-17 and GM-CSF, which promote the development of experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS). The protective role of STAT1 in MS and EAE has been largely attributed to its ability to limit pathogenic Th cells and promote Tregs. Using mice with selective deletion of STAT1 in T cells (STAT1CD4-Cre), we identified a potentially novel mechanism by which STAT1 regulates neuroinflammation independently of Foxp3+ Tregs. STAT1-deficient effector T cells became the target of NK cell–mediated killing, limiting their capacity to induce EAE. STAT1-deficient T cells promoted their own killing by producing more IL-2 that, in return, activated NK cells. Elimination of NK cells restored EAE susceptibility in STAT1CD4-Cre mice. Therefore, our study suggests that the STAT1 pathway can be manipulated to limit autoreactive T cells during autoimmunity directed against the CNS.

Identification of a distal enhancer that determines the expression pattern of acute phase marker C-reactive protein

C-reactive protein (CRP) is a major acute phase protein and inflammatory marker, the expression of which is largely liver specific and highly inducible. Enhancers are regulatory elements critical for the precise activation of gene expression, yet the contributions of enhancers to the expression pattern of CRP have not been well defined. Here, we identify a constitutively active enhancer (E1) located 37.7 kb upstream of the promoter of human CRP in hepatocytes. By using chromatin immunoprecipitation, luciferase reporter assay, in situ genetic manipulation, CRISPRi, and CRISPRa, we show that E1 is enriched in binding sites for transcription factors STAT3 and C/EBP-β and is essential for the full induction of human CRP during the acute phase. Moreover, we demonstrate that E1 orchestrates with the promoter of CRP to determine its varied expression across tissues and species through surveying activities of E1-promoter hybrids and the associated epigenetic modifications. These results thus suggest an intriguing mode of molecular evolution wherein expression-changing mutations in distal regulatory elements initiate subsequent functional selection involving coupling among distal/proximal regulatory mutations and activity-changing coding mutations.