Slow-and fast-twitch myofibers of adult skeletal muscles express unique sets of muscle-specific genes, and these distinctive programs of gene expression are controlled by variations in motor neuron activity. It is well established that, as a consequence of more frequent neural stimulation, slow fibers maintain higher levels of intracellular free calcium than fast fibers, but the mechanisms by which calcium may function as a messenger linking nerve activity to changes in gene expression in skeletal muscle have been unknown. Here, fiber-type-specific gene expression in skeletal muscles is shown to be controlled by a signaling pathway that involves calcineurin, a cyclosporin-sensitive, calcium-regulated serine/threonine phosphatase. Activation of calcineurin in skeletal myocytes selectively up-regulates slow-fiber-specific gene promoters. Conversely, inhibition of calcineurin activity by administration of cyclosporin A to intact animals promotes slow-to-fast fiber transformation. Transcriptional activation of slow-fiber-specific transcription appears to be mediated by a combinatorial mechanism involving proteins of the NFAT and MEF2 families. These results identify a molecular mechanism by which different patterns of motor nerve activity promote selective changes in gene expression to establish the specialized characteristics of slow and fast myofibers.

The transcription factor E-twenty-six related gene (ERG), which is overexpressed through gene fusion with the androgen-responsive gene transmembrane protease, serine 2 (TMPRSS2) in ∼40% of prostate tumors, is a key driver of prostate carcinogenesis. Ablation of ERG would disrupt a key oncogenic transcriptional circuit and could be a promising therapeutic strategy for prostate cancer treatment. Here, we show that ubiquitin-specific peptidase 9, X-linked (USP9X), a deubiquitinase enzyme, binds ERG in VCaP prostate cancer cells expressing TMPRSS2-ERG and deubiquitinates ERG in vitro. USP9X knockdown resulted in increased levels of ubiquitinated ERG and was coupled with depletion of ERG. Treatment with the USP9X inhibitor WP1130 resulted in ERG degradation both in vivo and in vitro, impaired the expression of genes enriched in ERG and prostate cancer relevant gene signatures in microarray analyses, and inhibited growth of ERG-positive tumors in three mouse xenograft models. Thus, we identified USP9X as a potential therapeutic target in prostate cancer cells and established WP1130 as a lead compound for the development of ERGdepleting drugs. P rostate cancer is the most common malignancy in men and the second or third leading cause of male cancer-related death in most Western countries, including the United States (1). Advanced prostate cancer initially responds to androgen ablation therapy, but hormone-refractory prostate cancer often times recurs, which has limited treatment options. Fusions of E-twentysix (ETS) transcription factor genes with androgen-responsive genes (2), mainly transmembrane protease, serine 2 (TMPRSS2), are present in up to 80% of prostate cancers. Patients with the most common ETS gene fusion TMPRSS2-ETS related gene (TMPRSS2-ERG) have a higher incidence of metastatic disease and cancer-related death compared with fusion-negative patients (3, 4), and in castration-resistant prostate cancer, TMPRSS2-ERG expression is frequently reactivated (5). In support of ERG being a key driver of prostate cancer, depletion of ERG by RNAi decreases proliferation and/or invasiveness in prostate cancer cell lines (2, 6), and ectopic expression of ERG in transgenic mice was shown to promote prostate oncogenesis in cooperation with the loss of tumor suppressors (7-12). TMPRSS2-driven overexpression of ERG controls a transcriptional network related to the development of prostate cancer and its progression to metastatic disease (13,14). This crucial role of ERG and the high incidence of the TMPRSS2-ERG gene fusion in prostate cancer have catapulted this protein into the forefront of new targets for therapeutic intervention (3,8). In the present study, we report the discovery of a deubiquitinase that stabilizes ERG in prostate cancer cells and demonstrate that pharmacological inhibition of this enzyme causes ERG depletion. Results USP9X is an ERG-Binding Protein.Proteins that interact with ERG in prostate cancer cells may modulate its activity, localization, or stability and could be harnessed as therapeutic target...

Abstract-Stem and progenitor cell populations occupy a specialized niche and are consequently exposed to hypoxic as well as oxidative stresses. We have previously established that the multidrug resistance protein Abcg2 is the molecular determinant of the side population (SP) progenitor cell population. We observed that the cardiac SP cells increase in number more than 3-fold within 3 days of injury. Transcriptome analysis of the SP cells isolated from the injured adult murine heart reveals increased expression of cytoprotective transcripts. Overexpression of Abcg2 results in an increased ability to consume hydrogen peroxide and is associated with increased levels of ␣-glutathione reductase protein expression. Importantly, overexpression of Abcg2 also conferred a cell survival benefit following exposure to hydrogen peroxide. To further examine the molecular regulation of the Abcg2 gene, we demonstrated that hypoxia-inducible factor (HIF)-2␣ binds an evolutionary conserved HIF-2␣ response element in the murine Abcg2 promoter. [1][2][3][4] This strategy defines a rare population of progenitor cells that can adopt alternative fates in permissive environments. [1][2][3] We have verified that the ability of SP cells to efflux Hoechst 33342 dye is dependent on the expression of Abcg2, which is a member of the family of ATP-binding cassette (ABC) transporters. 3,5,6 Although the functional role of the ABC transporters remains ill-defined, 7 we established that Abcg2 was able to confer the SP phenotype in a striated muscle cell line. 3 Stem and progenitor cell populations, including SP cells, are exposed to environmental stress by virtue of their physical location. Although oxidative stress attributable to unchecked levels of free radical-derived reactive oxygen species (ROS) can damage DNA, proteins, and lipids, 8 oxidative stress caused by modestly increased ROS can activate specific signal transduction pathways, leading to either senescence or apoptosis. 9 Previous transcriptome analyses of embryonic, hematopoietic, and neural stem cells revealed a common signature of gene expression in these stem cell populations. This profile includes transcripts that function as cytoprotective factors to provide resistance against environmental stress. 10 -12 Recent studies that examined circulating, bloodderived endothelial progenitor cells reveal enrichment for the expression of genes encoding for antioxidative factors that reduce sensitivity toward ROS-induced cell death. 13 Regulation of cytoprotective factors during injury states would be beneficial for survival and expansion of stem and progenitor cell populations.Members of the hypoxia-inducible factor (HIF) family are activated by multiple environmental stimuli. HIF-1␣, a master regulator for hypoxia-inducible gene expression, regulates gene expression to promote energy production as well as oxygen delivery in response to hypoxia. 14 -16 HIF-2␣, also known as endothelial PAS domain protein 1 (EPAS1), has many similarities with HIF-1␣. [17][18][19] However, several mole...

SUMMARY Efforts to identify and target glioblastoma (GBM) drivers have primarily focused on receptor tyrosine kinases (RTKs). Clinical benefits, however, have been elusive. Here, we identify a SRY-related box 2 (SOX2) transcriptional regulatory network that is independent of upstream RTKs and is capable of driving glioma initiating cells. We identified oligodendrocyte lineage transcription factor 2 (OLIG2) and zinc finger E-box binding homeobox 1 (ZEB1) as potential SOX2 targets, which are frequently co-expressed irrespective of driver mutations. In murine glioma models, we show that different combinations of tumor suppressor and oncogene mutations can activate Sox2, Olig2, and Zeb1 expression. We demonstrate that ectopic co-expression of the three transcription factors can transform tumor suppressor deficient astrocytes into glioma initiating cells in the absence of an upstream RTK oncogene. Finally, we demonstrate that the transcriptional inhibitor mithramycin downregulates SOX2 and its target genes, resulting in markedly reduced proliferation of GBM cells in vivo.