The transcriptional regulator Sin3A balances IL-17A and Foxp3 expression in primary CD4 T cells.

The Sin3 transcriptional regulator homolog A (Sin3A) is the core member of a multiprotein chromatin-modifying complex. Its inactivation at the CD4/CD8 double-negative stage halts further thymocyte development. Among various functions, Sin3A regulates STAT3 transcriptional activity, central to the differentiation of Th17 cells active in inflammatory disorders and opportunistic infections. To further investigate the consequences of conditional Sin3A inactivation in more mature precursors and post-thymic T cell, we have generated CD4-Cre and CD4-CreERT2 Sin3AF/F mice. Sin3A inactivation in vivo hinders both thymocyte development and peripheral T-cell survival. In vitro, in Th17 skewing conditions, Sin3A-deficient cells proliferate and acquire memory markers and yet fail to properly upregulate Il17a, Il23r, and Il22. Instead, IL-2+ and FOXP3+ are mostly enriched for, and their inhibition partially rescues IL-17A+ T cells. Notably, Sin3A deletion also causes an enrichment of genes implicated in the mTORC1 signaling pathway, overt STAT3 activation, and aberrant cytoplasmic RORγt accumulation. Thus, together our data unveil a previously unappreciated role for Sin3A in shaping critical signaling events central to the acquisition of immunoregulatory T-cell phenotypes.

The Transcriptional Regulator Sin3A Contributes to the Oncogenic Potential of STAT3.

Epigenetic silencing of promoter and enhancer regions is a common phenomenon in malignant cells. The transcription factor STAT3 is aberrantly activated in several tumors, where its constitutive acetylation accounts for the transcriptional repression of a number of tumor suppressor genes (TSG) via molecular mechanisms that remain to be understood. Using nucleophosmin-anaplastic lymphoma kinase-positive (NPM-ALK+) anaplastic large-cell lymphoma (ALCL) as model system, we found in cells and patient-derived tumor xenografts that STAT3 is constitutively acetylated as a result of ALK activity. STAT3 acetylation relied on intact ALK-induced PI3K- and mTORC1-dependent signaling and was sensitive to resveratrol. Resveratrol lowered STAT3 acetylation, rescued TSG expression, and induced ALCL apoptotic cell death. STAT3 constitutively bound the Sin3A transcriptional repressor complex, and both STAT3 and Sin3A bound the promoter region of silenced TSG via a resveratrol-sensitive mechanism. Silencing SIN3A caused reexpression of TSG, induced ALCL apoptotic cell death in vitro, and hindered ALCL tumorigenic potential in vivo. A constitutive STAT3-Sin3A interaction was also found in breast adenocarcinoma cells and proved critical for TSG silencing and cell survival. Collectively, these results suggest that oncogene-driven STAT3 acetylation and its constitutive association with Sin3A represent novel and concomitant events contributing to STAT3 oncogenic potential. SIGNIFICANCE: This study delineates the transcriptional regulatory complex Sin3A as a mediator of STAT3 transcriptional repressor activity and identifies the STAT3/Sin3A axis as a druggable target to antagonize STAT3-addicted tumors. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/79/12/3076/F1.large.jpg.See related commentary by Monteleone and Poli, p. 3031.

TYK2-induced phosphorylation of Y640 suppresses STAT3 transcriptional activity.

STAT3 is a pleiotropic transcription factor involved in homeostatic and host defense processes in the human body. It is activated by numerous cytokines and growth factors and generates a series of cellular effects. Of the STAT-mediated signal transduction pathways, STAT3 transcriptional control is best understood. Jak kinase dependent activation of STAT3 relies on Y705 phosphorylation triggering a conformational switch that is stabilized by intermolecular interactions between SH2 domains and the pY705 motif. We here show that a second tyrosine phosphorylation within the SH2 domain at position Y640, induced by Tyk2, negatively controls STAT3 activity. The Y640F mutation leads to stabilization of activated STAT3 homodimers, accelerated nuclear translocation and superior transcriptional activity following IL-6 and LIF stimulation. Moreover, it unlocks type I IFN-dependent STAT3 signalling in cells that are normally refractory to STAT3 transcriptional activation.

Targeting the tumor and its associated stroma: One and one can make three in adoptive T cell therapy of solid tumors.

Adoptive T cell therapy (ACT) has become a promising immunotherapeutic option for cancer patients. The proof for ACT therapeutic efficacy was first obtained with allogenic T cells and then reproduced with T cells isolated from patients' tumor samples (i.e. tumor-infiltrating lymphocytes). It is now clear that specificity of ACT products can be educated by genetically engineering T cells with classical T Cell Receptors (TCR) or chimeric antigen receptors (CAR). To date a poor accessibility of the tumor mass and a hostile microenvironment, influenced by genetic and epigenetic instability, mainly limit ACT therapeutic efficacy in the case of solid tumors. Available data indicate that these hurdles might be overcome by combinatorial therapeutic strategies targeting the tumor and its associated stroma. Here we review some of the available dual targeting strategies focusing on given combination of TCR/CAR-redirected T cell products and their association with drugs targeting the tumor-vessel and/or epigenetic modifiers, with the ability to sensitize tumors to T cell recognition. Existing data have proven synergistic effects in combined settings (one and one can indeed make three) and suggest that further benefit might be achieved by additional combinatorial therapeutic approaches (could one+one+one make ten?) in ACT of solid tumor.

The need for transparency and good practices in the qPCR literature.

Two surveys of over 1,700 publications whose authors use quantitative real-time PCR (qPCR) reveal a lack of transparent and comprehensive reporting of essential technical information. Reporting standards are significantly improved in publications that cite the Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines, although such publications are still vastly outnumbered by those that do not.

The HAT/HDAC interplay: multilevel control of STAT signaling.

Besides the transcription-promoting role of histone acetyltransferases (HATs) and the transcription-delimiting function of histone deacetylases (HDACs) through histone acetylation and deacetylation respectively, HATs and HDACs also regulate the activity of several non-histone proteins. This includes signal transducers and activators of transcription (STATs), key proteins in cytokine signaling. Unlike Tyr phosphorylation/dephosphorylation, which mainly acts as an on/off switch of STAT activity, the control exerted by HATs and HDACs appears multifaceted and far more complex than initially imagined. Our review focuses on the latest trends and novel hypotheses to explain differential context-dependent STAT regulation by complex posttranslational modification patterns. We chart the knowledge on how STATs interact with HATs and HDACs, and additionally bring a transcriptional regulatory and gene-set specific role for HDACs in the picture. Indeed, a growing amount of evidence demonstrates, paradoxically, that not only HAT but also HDAC activity can be required for STAT-dependent transcription, in a STAT subtype- and cell type-dependent manner. Referring to recent reports, we review and discuss the various molecular mechanisms that have recently been proposed to account for this peculiar regulation, in an attempt to shed more light on the difficult yet important question on how STAT specificity is being generated.

The Sin3a repressor complex is a master regulator of STAT transcriptional activity.

Tyrosine phosphorylation is a hallmark for activation of STAT proteins, but their transcriptional activity also depends on other secondary modifications. Type I IFNs can activate both the ISGF3 (STAT1:STAT2:IRF9) complex and STAT3, but with cell-specific, selective triggering of only the ISGF3 transcriptional program. Following a genome-wide RNAi screen, we identified the SIN3 transcription regulator homolog A (Sin3a) as an important mediator of this STAT3-targeted transcriptional repression. Sin3a directly interacts with STAT3 and promotes its deacetylation. SIN3A silencing results in a prolonged nuclear retention of activated STAT3 and enhances its recruitment to the SOCS3 promoter, concomitant with histone hyperacetylation and enhanced STAT3-dependent transcription. Conversely, Sin3a is required for ISGF3-dependent gene transcription and for an efficient IFN-mediated antiviral protection against influenza A and hepatitis C viruses. The Sin3a complex therefore acts as a context-dependent ISGF3/STAT3 transcriptional switch.

Opposed regulation of type I IFN-induced STAT3 and ISGF3 transcriptional activities by histone deacetylases (HDACS) 1 and 2.

The antiviral and antiproliferative responses mediated by type I interferons (IFNs) depend on JAK/STAT signaling and ISGF3 (STAT1:STAT2:IRF9)-dependent transcription. In addition, type I IFNs stimulate STAT3 activation in many cell types, an event generally associated with cell cycle progression, survival, and proliferation. To gather more insight into this functionally contradictive phenomenon, we studied the regulation of STAT3 transcriptional activity upon type I IFN treatment. We show that IFNα2 stimulation strongly induces STAT3 phosphorylation, nuclear translocation, and promoter binding, yet the activation of transcription of a STAT3-dependent reporter and endogenous genes, such as SOCS3 and c-FOS, is impaired. Simultaneous treatment with IFNα2 and trichostatin A, as well as combined HDAC1/HDAC2 silencing, restores STAT3-dependent reporter gene and endogenous gene expression, strongly suggesting that HDAC1 and HDAC2 are directly involved in repressing IFNα2-activated STAT3. Of note, single silencing of only one of the two HDACs does not lead to enhanced STAT3 activity, supporting a functional redundancy between these two enzymes. In sharp contrast, HDAC1 and HDAC2 activities are required for ISGF3-dependent gene expression. We conclude that HDAC1 and HDAC2 differentially modulate STAT activity in response to IFNα2: while they are required for the induction of ISGF3-responsive genes, they impair the transcription of STAT3-dependent genes.