In tomato (Solanum lycopersicum), as in other plants, the immunity hormone jasmonate (JA) triggers genome-wide transcriptional changes in response to pathogen and insect attack. These changes are largely regulated by the basic helix-loop-helix (bHLH) transcription factor MYC2. The function of MYC2 depends on its physical interaction with the MED25 subunit of the Mediator transcriptional coactivator complex. Although much has been learned about the MYC2-dependent transcriptional activation of JA-responsive genes, relatively less studied is the termination of JA-mediated transcriptional responses and the underlying mechanisms. Here, we report an unexpected function of MYC2 in regulating the termination of JA signaling through activating a small group of JA-inducible bHLH proteins, termed MYC2-TARGETED BHLH1 (MTB1), MTB2, and MTB3. MTB proteins negatively regulate JA-mediated transcriptional responses via their antagonistic effects on the functionality of the MYC2-MED25 transcriptional activation complex. MTB proteins impair the formation of the MYC2-MED25 complex and compete with MYC2 to bind to its target gene promoters. Therefore, MYC2 and MTB proteins form an autoregulatory negative feedback circuit to terminate JA signaling in a highly organized manner. We provide examples demonstrating that gene editing tools such as CRISPR/Cas9 open up new avenues to exploit MTB genes for crop protection.

BackgroundEndothelial E-selectin has been shown to play a pivotal role in mediating cell–cell interactions between breast cancer cells and endothelial monolayers during tumor cell metastasis. However, the counterreceptor for E-selectin and its role in mediating breast cancer cell transendothelial migration remain unknown.Methodology/Principal FindingsBy assessing migration of various breast cancer cells across TNF-α pre-activated human umbilical vein endothelial cells (HUVECs), we found that breast cancer cells migrated across HUVEC monolayers differentially and that transmigration was E-selectin dependent. Cell surface labeling with the E-selectin extracellular domain/Fc chimera (exE-selectin/Fc) showed that the transmigration capacity of breast cancer cells was correlated to both the expression level and localization pattern of E-selectin binding protein(s) on the tumor cell surface. The exE-selectin/Fc strongly bound to metastatic MDA-MB-231, MDA-MB-435 and MDA-MB-468 cells, but not non-metastatic MCF-7 and T47D cells. Binding of exE-selectin/Fc was abolished by removal of tumor cell surface sialyl lewis x (sLex) moieties. Employing an exE-selectin/Fc affinity column, we further purified the counterreceptor of E-selectin from metastatic breast cancer cells. The N-terminal protein sequence and cDNA sequence identified this E-selectin ligand as a ∼170 kD human CD44 variant 4 (CD44v4). Purified CD44v4 showed a high affinity for E-selectin via sLex moieties and, as expected, MDA-MB-231 cell adhesion to and migration across HUVEC monolayers were significantly reduced by down-regulation of tumor cell CD44v4 via CD44v4-specific siRNA.Conclusions/SignificanceWe demonstrated, for the first time, that breast cancer cell CD44v4 is a major E-selectin ligand in facilitating tumor cell migration across endothelial monolayers. This finding offers new insights into the molecular basis of E-selectin–dependent adhesive interactions that mediate breast cancer cell transendothelial metastasis.

Chronic inflammation underscores the pathogenesis of a range of human diseases. Lipopolysaccharide (LPS) elicits strong pro-inflammatory response in macrophages via the transcription factor NF-κB. The epigenetic mechanism underlying LPS-induced pro-inflammatory transcription is not completely appreciated. Herein we describe a role for myocardin related transcription factor A, or MRTF-A, in this process. MRTF-A over-expression potentiated while MRTF-A silencing dampened NF-κB dependent pro-inflammatory transcription. MRTF-A deficiency also alleviated the synthesis of pro-inflammatory mediators in a mouse model of colitis. LPS promoted the recruitment of MRTF-A to the promoters of pro-inflammatory genes in a NF-κB dependent manner. Reciprocally, MRTF-A influenced the nuclear enrichment and target binding of NF-κB. Mechanistically, MRTF-A was necessary for the accumulation of active histone modifications on NF-κB target promoters by communicating with the histone H3K4 methyltransferase complex (COMPASS). Silencing of individual members of COMPASS, including ASH2, WDR5, and SET1, down-regulated the production of pro-inflammatory mediators and impaired the NF-κB kinetics. In summary, our work has uncovered a previously unknown function for MRTF-A and provided insights into the rationalized development of anti-inflammatory therapeutic strategies.

Diabetic nephropathy (DN) is one of the most common complications associated with diabetes and characterized by renal microvascular injury along with accelerated synthesis of extracellular matrix proteins causing tubulointerstitial fibrosis. Production of type I collagen, the major component of extracellular matrix, is augmented during renal fibrosis after chronic exposure to hyperglycemia. However, the transcriptional modulator responsible for the epigenetic manipulation leading to induction of type I collagen genes is not clearly defined. We show here that tubulointerstitial fibrosis as a result of DN was diminished in myocardin-related transcription factor A (MRTF-A) -deficient mice. In cultured renal tubular epithelial cells and the kidneys of mice with DN, MRTF-A was induced by glucose and synergized with glucose to activate collagen transcription. Notably, MRTF-A silencing led to the disappearance of prominent histone modifications indicative of transcriptional activation, including acetylated histone H3K18/K27 and trimethylated histone H3K4. Detailed analysis revealed that MRTF-A recruited p300, a histone acetyltransferase, and WD repeat-containing protein 5 (WDR5), a key component of the histone H3K4 methyltransferase complex, to the collagen promoters and engaged these proteins in transcriptional activation. Estradiol suppressed collagen production by dampening the expression and binding activity of MRTF-A and interfering with the interaction between p300 and WDR5 in renal epithelial cells. Therefore, targeting the MRTF-A-associated epigenetic machinery might yield interventional strategies against DN-associated renal fibrosis.

-Excessive accumulation of reactive oxygen species (ROS), catalyzed by the NADPH oxidases (NOX), is involved in the pathogenesis of ischemia-reperfusion (I/R) injury. The underlying epigenetic mechanism remains elusive. -We evaluated the potential role of megakaryocytic leukemia 1, or MKL1, as a bridge linking epigenetic activation of NOX to ROS production and cardiac ischemia-reperfusion injury. -Following I/R injury, MKL1 deficient (KO) mice exhibited smaller myocardial infarction along with improved heart function compared to wild type (WT) littermates. Similarly, pharmaceutical inhibition of MKL1 with CCG-1423 also attenuated myocardial infarction and improved heart function in mice. Amelioration of I/R injury as a result of MKL1 deletion or inhibition was accompanied by reduced ROS and In response to I/R, MKL1 levels were specifically elevated in macrophages, but not in cardiomyocytes, in the heart. Of note, macrophage-specific deletion (MφcKO), instead of cardiomyocyte-restricted ablation (CMcKO), of MKL1 in mice led to similar improvements of infarct size, heart function, and myocardial ROS generation. Reporter assay and ChIP assay revealed that MKL1 directly bound to the promoters of NADPH oxidase (NOX) genes to activate NOX transcription. Mechanistically, MKL1 recruited the histone acetyltransferase MOF to modify the chromatin structure surrounding the NOX promoters. Knockdown of MOF in macrophages blocked hyoxia/re-oxygenation-induced NOX transactivation and ROS accumulation. Of importance, pharmaceutical inhibition of MOF with MG-149 significantly down-regulated NOX1/NOX4 expression, dampened ROS production, and normalized myocardial function in mice exposed to I/R injury. Finally, administration of a specific NOX1/4 inhibitor GKT137831 dampened ROS generation and rescued heart function following I/R in mice. -Our data delineate an MKL1-MOF-NOX axis in macrophages that contributes to I/R injury and as such have provided novel therapeutic targets in the treatment of ischemic heart disease.

Rationale Endothelial dysfunction inflicted by inflammation is found in a host of cardiovascular pathologies. One hallmark event in this process is the aggregation and adhesion of leukocyte to the vessel wall mediated by the up-regulation of adhesion molecules (CAM) in endothelial cells at the transcriptional level. The epigenetic modulator(s) of CAM transactivation and its underlying pathophysiological relevance remain poorly defined. Objective Our goal was to determine the involvement of Brg1 and Brm in CAM transactivation and its relevance in the pathogenesis of atherosclerosis. Methods and Results In the present study, we report that pro-inflammatory stimuli augmented the expression of Brg1 and Brm in vitro in cultured endothelial cells and in vivo in arteries isolated from rodents. Over-expression of Brg1 and Brm promoted whereas knockdown of Brg1 and Brm abrogated transactivation of adhesion molecules and leukocyte adhesion induced by inflammatory signals. Brg1 and Brm interacted with and were recruited to the CAM promoters by NF-κB/p65. Conversely, depletion of Brg1 and Brm disrupted the kinetics of p65 binding on CAM promoters and crippled CAM activation. Silencing of Brg1 and Brm also altered key epigenetic changes associated with CAM transactivation. Of intrigue, 17β-estradiol antagonized both the expression and activity of Brg1/Brm. Most importantly, endothelial-targeted elimination of Brg1/Brm conferred atheroprotective effects to Apoe−/− mice on a Western diet. Conclusion Therefore, our data suggest that Brg1 and Brm integrate various pro-inflammatory cues into CAM transactivation and endothelial malfunction and as such may serve as potential therapeutic targets in treating inflammation related cardiovascular diseases.

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Myocardial infarction (MI) dampens heart function and poses a great health risk. The class III deacetylase sirtuin 1 (SIRT1) is known to confer cardioprotection. SIRT1 expression is downregulated in the heart by a number of stress stimuli that collectively drive the pathogenesis of MI, although the underlying mechanism remains largely obscure. Here we show that in primary rat neonatal ventricular myocytes (NRVMs), ischaemic or oxidative stress leads to a rapid upregulation of SUV39H, the mammalian histone H3K9 methyltransferase, paralleling SIRT1 downregulation. Compared to wild-type littermates, SUV39H knockout mice are protected from MI. Likewise, suppression of SUV39H activity with chaetocin attenuates cardiac injury following MI. Mechanistically, SUV39H cooperates with heterochromatin protein 1 gamma (HP1γ) to catalyse H3K9 trimethylation on the SIRT1 promoter and represses SIRT1 transcription. SUV39H augments intracellular ROS levels in a SIRT1-dependent manner. Our data identify a previously unrecognized role for SUV39H linking SIRT1 trans-repression to myocardial infarction.