STAT3 Protein Regulates Vascular Smooth Muscle Cell Phenotypic Switch by Interaction with Myocardin.

The JAK-STAT3 signaling pathway is one of the critical pathways regulating cell proliferation, differentiation, and apoptosis. Myocardin is regarded as a key mediator for the change of smooth muscle phenotypes. However, the relationship between STAT3 and myocardin in the vascular smooth muscle cell (VSMC) phenotypic switch has not been investigated. The goal of this study was to investigate the molecular mechanism by which STAT3 affects the myocardin-regulated VSMC phenotypic switch. Data presented in this study demonstrated that STAT3 was rapidly up-regulated after stimulation with VEGF. Inhibition of the STAT3 activation process impaired VSMC proliferation and enhanced the expression of VSMC contractile genes by increasing serum-response factor binding to the CArG-containing regions of VSMC-specific contractile genes. In contrast, the interaction between serum-response factor and its co-activator myocardin was reduced by overexpression of STAT3. In addition, treated VEGF inhibited the transcription activity of myocardin, and overexpression of STAT3 inhibited myocardin-induced up-regulation of VSMC contractile phenotype-specific genes. Although myocardin and STAT3 are negatively correlated, interestingly, both of them can enhance the expression of VEGF, suggesting a feedback loop to regulate the VSMC phenotypic switch. Taken together, these results indicate that the JAK-STAT3 signaling pathway plays a key role in controlling the phenotypic switch of VSMCs through the interactions between STAT3 and myocardin by various coordinated gene regulation pathways and feedback loops.

Myocardin inhibits estrogen receptor alpha-mediated proliferation of human breast cancer MCF-7 cells via regulating MicroRNA expression.

Myocardin is frequently repressed during human malignant transformation, and restoration of myocardin expression in sarcoma cells contributes to the inhibition of malignant growth. However, its role in breast carcinoma has barely been addressed. Here, we reported that myocardin could inhibit the proliferation of MCF-7 cells. Notably, we show that myocardin inhibited ERα-mediated proliferation of breast cancer MCF-7 via impairing ER-dependent transcriptional activation, mainly through the inhibition of the activity of ERα. Importantly, the molecular mechanism for the inhibition of the ERα-mediated proliferation is that myocardin inhibited the transcription and expression of ERα-induced PCNA, Ki-67, and E2F1 to impair ERα-mediated proliferation of breast cancer MCF-7. Interestingly, myocardin significantly enhanced the transcription and expression of miR-885 depending on the CArG box in miR-885 promoter, and miR-885 targeted the 3' untranslated regions (UTR) of E2F1 to silence the expression of E2F1. Thus, our data provided important and novel insights into how myocardin may deeply influence ERα-mediated breast cancer proliferation. In conclusion, myocardin could be seen as a breast cancer tumor suppressor so that it will provide new ideas for the treatment of breast cancer. © 2016 IUBMB Life, 68(6):477-487, 2016.

MRTF-A and STAT3 promote MDA-MB-231 cell migration via hypermethylating BRSM1.

Breast cancer is the leading cause of cancer death in women worldwide which is closely related to metastasis. But the exact molecular mechanism of metastasis is still not fully understood. We now report that both MRTF-A and STAT3 play important roles in migration of MDA-MB-231 breast cancer cells. Moreover, MRTF-A and STAT3 synergistically increased MDA-MB-231 cell migration by promoting the expression of migration markers urokinase-type plasminogen activator (uPA) and osteopontin (OPN) and inhibiting the expression of breast cancer metastasis suppressor 1 (BRMS1). Luciferase reporter assays demonstrated that MRTF-A and STAT3 do not affect transcription of the BRMS1 promoter. Instead, we identified a newly molecular mechanism by which MRTF-A and STAT3 synergistically controlled MDA-MB-231 cell migration by recruiting DNMT1 to hypermethylate the promoter of BRMS1 and thus affect the expression of BRMS1. Interestingly, physical interaction between MRTF-A and STAT3 synergistically promotes the transactivity of DNMT1 by binding to the GAS element within the DNMT1 promoter. Our data thus provide important and novel insights into the roles of MRTF-A and STAT3 in regulating MDA-MB-231 cell migration.

VEGF-A Stimulates STAT3 Activity via Nitrosylation of Myocardin to Regulate the Expression of Vascular Smooth Muscle Cell Differentiation Markers.

Vascular endothelial growth factor A (VEGF-A) is a pivotal player in angiogenesis. It is capable of influencing such cellular processes as tubulogenesis and vascular smooth muscle cell (VSMC) proliferation, yet very little is known about the actual signaling events that mediate VEGF-A induced VSMC phenotypic switch. In this report, we describe the identification of an intricate VEGF-A-induced signaling cascade that involves VEGFR2, STAT3, and Myocardin. We demonstrate that VEGF-A promotes VSMC proliferation via VEGFR2/STAT3-mediated upregulating the proliferation of markers like Cyclin D1 and PCNA. Specifically, VEGF-A leads to nitrosylation of Myocardin, weakens its effect on promoting the expression of contractile markers and is unable to inhibit the activation of STAT3. These observations reinforce the importance of nitric oxide and S-nitrosylation in angiogenesis and provide a mechanistic pathway for VEGF-A-induced VSMC phenotypic switch. In addition, Myocardin, GSNOR and GSNO can create a negative feedback loop to regulate the VSMC phenotypic switch. Thus, the discovery of this interactive network of signaling pathways provides novel and unexpected therapeutic targets for angiogenesis-dependent diseases.

STAT3 regulated ATR via microRNA-383 to control DNA damage to affect apoptosis in A431 cells.

Skin cancer is a major cause of morbidity and mortality worldwide. Mounting evidence shows that exposure of the skin to solar UV radiation results in inflammation, oxidative stress, DNA damage, dysregulation of cellular signaling pathways and immunosuppression thereby resulting in skin cancer. Signal transducer and activator of transcription 3 (STAT3) is well known to function as an anti-apoptotic factor, especially in numerous malignancies, but the relationship between STAT3 activation and DNA damage response in skin cancer is still not fully understood. We now report that STAT3 inhibited DNA damage induced by UV and STAT3 mediated upregulation of GADD45γ and MDC-1 and the phosphorylation of H2AX in UV induced DNA damage. Notably, STAT3 can increase the expression of ATR in A431 cells. Luciferase assay shows that STAT3 activates the transcription of ATR promoter. More importantly, microRNA-383 suppressed ATR expression by targeting 3' (untranslated regions)UTR of ATR in A431 cells, and STAT3 down-regulates the transcription of miR-383 promoter. Thus, these results reveal the new insight that ATR is down-regulated by STAT3-regulated microRNA-383 in A431 cells. Moreover, overexpression of STAT3 enhanced expression of antiapoptosis genes BCL-1 and MCL-1, and depletion of STAT3 sensitized A431 cells to apoptotic cell death following UV. Collectively, these studies suggest that STAT3 may be a potential target for both the prevention and treatment of human skin cancer.

NF-κB (p65) negatively regulates myocardin-induced cardiomyocyte hypertrophy through multiple mechanisms.

Myocardin is well known to play a key role in the development of cardiomyocyte hypertrophy. But the exact molecular mechanism regulating myocardin stability and transactivity to affect cardiomyocyte hypertrophy has not been studied clearly. We now report that NF-κB (p65) can inhibit myocardin-induced cardiomyocyte hypertrophy. Then we explore the molecular mechanism of this response. First, we show that p65 can functionally repress myocardin transcriptional activity and also reduce the protein expression of myocardin. Second, the function of myocardin can be regulated by epigenetic modifications. Myocardin sumoylation is known to transactivate cardiac genes, but whether p65 can inhibit SUMO modification of myocardin is still not clear. Our data show that p65 weakens myocardin transcriptional activity through attenuating SUMO modification of myocardin by SUMO1/PIAS1, thereby impairing myocardin-mediated cardiomyocyte hypertrophy. Furthermore, the expression of myocardin can be regulated by several microRNAs, which play important roles in the development and function of the heart and muscle. We next investigated potential role of miR-1 in cardiac hypotrophy. Our results show that p65 can upregulate the level of miR-1 and miR-1 can decrease protein expression of myocardin in cardiac myocytes. Notably, miR-1 expression is also controlled by myocardin, leading to a feedback loop. These data thus provide important and novel insights into the function that p65 inhibits myocardin-mediated cardiomyocyte hypertrophy by downregulating the expression and SUMO modification of myocardin and enhancing the expression of miR-1.