The CREB coactivator TORC2 functions as a calcium- and cAMP-sensitive coincidence detector

Elevations in circulating glucose and gut hormones during feeding promote pancreatic islet cell viability in part via the calcium- and cAMP-dependent activation of the transcription factor CREB. Here, we describe a signaling module that mediates the synergistic effects of these pathways on cellular gene expression by stimulating the dephosphorylation and nuclear entry of TORC2, a CREB coactivator. This module consists of the calcium-regulated phosphatase calcineurin and the Ser/Thr kinase SIK2, both of which associate with TORC2. Under resting conditions, TORC2 is sequestered in the cytoplasm via a phosphorylation-dependent interaction with 14-3-3 proteins. Triggering of the calcium and cAMP second messenger pathways by glucose and gut hormones disrupts TORC2:14-3-3 complexes via complementary effects on TORC2 dephosphorylation; calcium influx increases calcineurin activity, whereas cAMP inhibits SIK2 kinase activity. Our results illustrate how a phosphatase/kinase module connects two signaling pathways in response to nutrient and hormonal cues

Activation of Crtc2/Creb1 in skeletal muscle enhances weight loss during intermittent fasting

The Creb-Regulated Transcriptional Coactivator (Crtc) family of transcriptional coregulators drive Creb1-mediated transcription effects on metabolism in many tissues, but the in vivo effects of Crtc2/Creb1 transcription on skeletal muscle metabolism are not known. Skeletal muscle-specific overexpression of Crtc2 (Crtc2 mice) induced greater mitochondrial activity, metabolic flux capacity for both carbohydrates and fats, improved glucose tolerance and insulin sensitivity, and increased oxidative capacity, supported by upregulation of key metabolic genes. Crtc2 overexpression led to greater weight loss during alternate day fasting (ADF), selective loss of fat rather than lean mass, maintenance of higher energy expenditure during the fast and reduced binge-eating during the feeding period. ADF downregulated most of the mitochondrial electron transport genes, and other regulators of mitochondrial function, that were substantially reversed by Crtc2-driven transcription. Glucocorticoids acted with AMPK to drive atrophy and mitophagy, which was reversed by Crtc2/Creb1 signaling. Crtc2/Creb1-mediated signaling coordinates metabolic adaptations in skeletal muscle that explain how Crtc2/Creb1 regulates metabolism and weight loss

Striatal microRNA controls cocaine intake through CREB signaling

Cocaine addiction is characterized by a gradual loss of control over drug use, but molecular mechanisms regulating vulnerability to this process remain unclear. Here we report that microRNA-212 (miR-212) is upregulated in the dorsal striatum of rats with a history of extended access to cocaine. Striatal miR-212 decreases responsiveness to the motivational properties of cocaine by dramatically amplifying the stimulatory effects of the drug on CREB signaling. This action occurs through miR-212-enhanced Raf-1 activity, resulting in adenylyl cyclase sensitization and increased expression of the essential CREB co-activator TORC (Transducer of Regulated CREB; also known as CRTC). Our findings suggest that striatal miR-212 signaling plays a key role in determining vulnerability to cocaine addiction, reveal novel molecular regulators that control the complex actions of cocaine in brain reward circuitries, and provide an entirely new direction for the development of anti-addiction therapeutics based on modulation of noncoding RNAs

Bipartite functions of the CREB co-activators selectively direct alternative splicing or transcriptional activation

The CREB regulated transcription co-activators (CRTCs) regulate many biological processes by integrating and converting environmental inputs into transcriptional responses. Although the mechanisms by which CRTCs sense cellular signals are characterized, little is known regarding how CRTCs contribute to the regulation of cAMP inducible genes. Here we show that these dynamic regulators, unlike other co-activators, independently direct either pre-mRNA splice-site selection or transcriptional activation depending on the cell type or promoter context. Moreover, in other scenarios, the CRTC co-activators coordinately regulate transcription and splicing. Mutational analyses showed that CRTCs possess distinct functional domains responsible for regulating either pre-mRNA splicing or transcriptional activation. Interestingly, the CRTC1–MAML2 oncoprotein lacks the splicing domain and is incapable of altering splice-site selection despite robustly activating transcription. The differential usage of these distinct domains allows CRTCs to selectively mediate multiple facets of gene regulation, indicating that co-activators are not solely restricted to coordinating alternative splicing with increase in transcriptional activity

Metformin Inhibits Follicle-Stimulating Hormone (FSH) Action in Human Granulosa Cells: Relevance to Polycystic Ovary Syndrome

Background: Women with anovulatory polycystic ovary syndrome (PCOS) are generally insulin-resistant and as a consequence are often treated with the biguanide metformin. Results with metformin have, however, been variable with some studies demonstrating induction of regular cycles and an increase in ovulation, whereas others do not. Hence more understanding is needed regarding the mechanism of metformin's actions in ovarian granulosa cells especially in light of previous demonstrations of direct actions. Objective: The aim of this study was to investigate metformin's interaction with the FSH/cAMP/protein kinase A pathway, which is the primary signaling pathway controlling CYP19A1 (aromatase) expression in the ovary. Methods: The effect of metformin on FSH and forskolin-stimulated aromatase expression in human granulosa cells was measured by quantitative real-time PCR. Activity was assessed after transfection with a promoter II-luciferase construct, and by an RIA measuring conversion of androgen to estrogens. The effect on FSH receptor (FSHR) mRNA was assessed by quantitative PCR. Levels of phosphorylated cAMP response element binding protein (CREB) and CREB-regulated transcription coactivator 2 (CRTC2) were measured by Western blotting and cAMP by a bioluminescent assay. Results: Metformin markedly reduced FSH but not forskolin-stimulated aromatase expression and activity. This effect was exerted by inhibition of basal and ligand-induced up-regulation of FSHR expression. Metformin also reduced FSH-induced phosphorylation of CREB and hence CRE activity, which could potentially disrupt the CREB–CREB-binding protein–CRTC2 coactivator complex that binds to CRE in promoter II of the aromatase gene. This is mediated in an AMP-activated protein kinase-independent manner, and does not involve alteration of cAMP levels. Conclusion: These finding have implications for the use of metformin in the treatment of anovulation in women with PCOS