The Janus kinase inhibitor (baricitinib) suppresses the rheumatoid arthritis active marker gliostatin/thymidine phosphorylase in human fibroblast-like synoviocytes

Gliostatin/thymidine phosphorylase (GLS/TP) is known to have angiogenic and arthritogenic activities in the pathogenesis of rheumatoid arthritis (RA). The novel oral Janus kinase (JAK) inhibitor baricitinib has demonstrated high efficacy in RA. However, the effect of baricitinib on fibroblast-like synoviocytes (FLSs), a key component of invasive synovitis, has not been still elucidated. This study investigated whether GLS/TP production could be regulated by JAK/signal transducers and activators of transcription (STAT) signaling in FLSs derived from patients with RA. FLSs were cultured and stimulated by interferon (IFN)γ in the presence of baricitinib. Expression levels of GLS/TP were determined using reverse transcription-polymerase chain reaction (RT-PCR), enzyme-linked immunosorbent assay (ELISA), and immunocytochemistry. Phosphorylation of STAT proteins was investigated by Western blot. In cultured FLSs, GLS/TP mRNA and protein levels were significantly induced by treatment with IFNγ and these inductions were suppressed by baricitinib treatment. Baricitinib inhibited IFNγ-induced STAT1 phosphorylation, while JAK/STAT activation played a pivotal role in IFNγ-mediated GLS/TP upregulation in RA. These results suggested that baricitinib suppressed IFNγ-induced GLS/TP expression by inhibiting JAK/STAT signaling, resulting in the attenuation of neovascularization, synovial inflammation, and cartilage destruction.

Mechanisms of Viral Degradation of Cellular Signal Transducer and Activator of Transcription 2

Virus infection of eukaryotes triggers cellular innate immune response, a major arm of which is the type I interferon (IFN) family of cytokines. Binding of IFN to cell surface receptors triggers a signaling cascade in which the signal transducer and activator of transcription 2 (STAT2) plays a key role, ultimately leading to an antiviral state of the cell. In retaliation, many viruses counteract the immune response, often by the destruction and/or inactivation of STAT2, promoted by specific viral proteins that do not possess protease activities of their own. This review offers a summary of viral mechanisms of STAT2 subversion with emphasis on degradation. Some viruses also destroy STAT1, another major member of the STAT family, but most viruses are selective in targeting either STAT2 or STAT1. Interestingly, degradation of STAT2 by a few viruses requires the presence of both STAT proteins. Available evidence suggests a mechanism in which multiple sites and domains of STAT2 are required for engagement and degradation by a multi-subunit degradative complex, comprising viral and cellular proteins, including the ubiquitin–proteasomal system. However, the exact molecular nature of this complex and the alternative degradation mechanisms remain largely unknown, as critically presented here with prospective directions of future study.

Tetramerization of STAT5 promotes autoimmune-mediated neuroinflammation

Signal tranducer and activator of transcription 5 (STAT5) plays a critical role in mediating cellular responses following cytokine stimulation. STAT proteins critically signal via the formation of dimers, but additionally, STAT tetramers serve key biological roles, and we previously reported their importance in T and natural killer (NK) cell biology. However, the role of STAT5 tetramerization in autoimmune-mediated neuroinflammation has not been investigated. Using the STAT5 tetramer-deficient Stat5a-Stat5b N-domain double knockin (DKI) mouse strain, we report here that STAT5 tetramers promote the pathogenesis of experimental autoimmune encephalomyelitis (EAE). The mild EAE phenotype observed in DKI mice correlates with the impaired extravasation of pathogenic T-helper 17 (Th17) cells and interactions between Th17 cells and monocyte-derived cells (MDCs) in the meninges. We further demonstrate that granulocyte-macrophage colony-stimulating factor (GM-CSF)–mediated STAT5 tetramerization regulates the production of CCL17 by MDCs. Importantly, CCL17 can partially restore the pathogenicity of DKI Th17 cells, and this is dependent on the activity of the integrin VLA-4. Thus, our study reveals a GM-CSF-STAT5 tetramer-CCL17 pathway in MDCs that promotes autoimmune neuroinflammation.

Simultaneous assessment of eight phosphorylated STAT residues in T-cells by flow cytometry.

Signal transducer and activator of transcription (STAT) proteins are a family of transcription factors controlling functions in immune responses and other cell types. Given their importance, we developed a flow cytometry panel to assess eight phosphorylated STAT residues in human T-cells, including six tyrosine residues across six STAT proteins (STAT1, STAT2, STAT3, STAT4, STAT5a, STAT6) and additional serine residues on STAT1 and STAT3. We applied this protocol to test the in vitro induction of pSTATs in response to CD3/CD28 activation and a panel of recombinant cytokines. We also assessed the pSTAT expression profiles of naive CD4+ T-cells polarized to Th1, Th2, Th17 or iTregs. pSTAT1(S727), pSTAT2(Y689) and pSTAT3(S727) were constitutively expressed in most T-cells, even in the absence of stimulation. For pSTAT1(S727) and pSTAT3(S727), we observed two positive states, high and low. Conversely, expression of pSTAT1(Y701), pSTAT3(Y705), pSTAT4(Y693) and pSTAT6(Y641) were absent in resting T-cells and only expressed with CD3/CD28 activation or with specific cytokines. Variable frequencies of pSTAT5a(Y694) expression were observed in resting T-cells, which increased with activation or specific cytokine stimulation (e.g. IL-2). IFN-beta stimulation enhanced frequencies of expressing cells for all pSTATs. Correlations among several pSTATs, particularly pSTAT1(S727)high and pSTAT3(S727)high were observed. While polarization resulted in increases in canonically associated pSTATs, other non-canonical pSTAT changes were also observed. Collectively, we developed, optimized, and tested a sensitive and rapid approach for simultaneously assessing phosphorylation of six STAT proteins. Using this approach, we made several novel observations of T-cell pSTAT induction in response to stimuli.

STAT proteins: a kaleidoscope of canonical and non-canonical functions in immunity and cancer

STAT proteins represent an important family of evolutionarily conserved transcription factors that play key roles in diverse biological processes, notably including blood and immune cell development and function. Classically, STAT proteins have been viewed as inducible activators of transcription that mediate cellular responses to extracellular signals, particularly cytokines. In this ‘canonical’ paradigm, latent STAT proteins become tyrosine phosphorylated following receptor activation, typically via downstream JAK proteins, facilitating their dimerization and translocation into the nucleus where they bind to specific sequences in the regulatory region of target genes to activate transcription. However, growing evidence has challenged this paradigm and identified alternate ‘non-canonical’ functions, such as transcriptional repression and roles outside the nucleus, with both phosphorylated and unphosphorylated STATs involved. This review provides a revised framework for understanding the diverse kaleidoscope of STAT protein functional modalities. It further discusses the implications of this framework for our understanding of STAT proteins in normal blood and immune cell biology and diseases such as cancer, and also provides an evolutionary context to place the origins of these alternative functional modalities.

Impact of STAT Proteins in Tumor Progress and Therapy Resistance in Advanced and Metastasized Prostate Cancer

Signal transducers and activators of transcription (STATs) are a family of transcription factors involved in several biological processes such as immune response, cell survival, and cell growth. However, they have also been implicated in the development and progression of several cancers, including prostate cancer (PCa). Although the members of the STAT protein family are structurally similar, they convey different functions in PCa. STAT1, STAT3, and STAT5 are associated with therapy resistance. STAT1 and STAT3 are involved in docetaxel resistance, while STAT3 and STAT5 are involved in antiandrogen resistance. Expression of STAT3 and STAT5 is increased in PCa metastases, and together with STAT6, they play a crucial role in PCa metastasis. Further, expression of STAT3, STAT5, and STAT6 was elevated in advanced and high-grade PCa. STAT2 and STAT4 are currently less researched in PCa. Since STATs are widely involved in PCa, they serve as potential therapeutic targets. Several inhibitors interfering with STATs signaling have been tested unsuccessfully in PCa clinical trials. This review focuses on the respective roles of the STAT family members in PCa, especially in metastatic disease and provides an overview of STAT-inhibitors evaluated in clinical trials.

A strategy to suppress STAT1 signalling conserved in pathogenic poxviruses and paramyxoviruses

The induction of interferon-stimulated genes by signal transducer and activator of transcription (STAT) proteins, is a critical host defence to fight virus infections. Here, a highly expressed poxvirus protein 018 is shown to inhibit IFN-induced signalling by binding the SH2 domain of STAT1 to prevent STAT1 association with an activated IFN receptor. Despite the presence of additional inhibitors of IFN-induced signalling, a poxvirus lacking 018 was attenuated in mice. The 2.0-angstrom crystal structure of the 018:STAT1 complex reveals a mechanism for a high-affinity, pTyr-independent mode of binding to an SH2 domain. Furthermore, the STAT1 binding motif of 018 shows sequence similarity to the STAT1-binding proteins from Nipah virus, which like 018, block the association of STAT1 with an IFN receptor. Taken together, these results provide detailed mechanistic insight into a potent mode of STAT1 antagonism, found to exist in genetically diverse virus families.

The Many Faces of JAKs and STATs Within the COVID-19 Storm

The positive-sense single stranded RNA virus, Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), resulted in a global pandemic with horrendous health and economic consequences not seen in a century. At a finer scale, immunologically, many of these devastating effects by SARS-CoV-2 can be traced to a “cytokine storm” resulting in the simultaneous activation of Janus Kinases (JAKs) and Signal Transducers and Activators of Transcription (STAT) proteins downstream of the many cytokine receptor families triggered by elevated cytokines found in Coronavirus Disease 2019 (COVID-19). In this report, cytokines found in the storm are discussed in relation to the JAK-STAT pathway in response to SARS-CoV-2 and the lessons learned from RNA viruses and previous Coronaviruses (CoVs). Therapeutic strategies to counteract the SARS-CoV-2 mediated storm are discussed with an emphasis on cell signaling and JAK inhibition.

Altered expression of STAT genes in periodontitis

Signal Transducer and Activator of Transcription (STAT) pathway is functionally located downstream of Janus kinases proteins and can integrate signals from diverse pathways, thus regulating several aspects of immune responses. Although contribution of STAT proteins in the pathogenesis of several inflammatory conditions has been confirmed, their role in the development of periodontitis has been less appraised. Thus, we assessed levels of STAT transcripts in the periodontal tissues and circulation of affected individuals compared with the corresponding controls. Expression of STAT1 was remarkably lower in tissues samples of patients compared with control tissues (Ratio of mean expression (RME) = 0.15, SE = 0.99, P value = 0.01). Expression of STAT3 was lower in total periodontitis tissues compared with total control tissues (RME = 0.20, SE = 0.95, P value = 0.02). Expression of STAT6 was higher in total periodontitis tissues compared with total control tissues (RME = 0.5.38, SE = 0.74, P value < 0.001). Expressions of other STAT genes were statistically similar in tissues obtained from cases and controls. Moreover, blood levels of all STAT genes were statistically similar between patients and controls. Correlation analysis demonstrated significant correlations between tissues levels of individual STAT genes as well as between their blood levels. However, tissue and blood levels of each STAT gene were not correlated. The current investigation potentiates the role of certain STAT genes in the development of this immune-related condition and warrants functional assays to clarify the mechanism.