Response Element-binding Protein Signaling Research Articles

Developmental transcriptomic analyses for mechanistic insights into critical pathways involved in embryogenesis of pelagic mahi-mahi (Coryphaena hippurus).

Mahi-mahi (Coryphaena hippurus) is a commercially and ecologically important species of fish occurring in tropical and temperate waters worldwide. Understanding early life events is crucial for predicting effects of environmental stress, which is largely restricted by a lack of genetic resources regarding expression of early developmental genes and regulation of pathways. The need for anchoring developmental stages to transcriptional activities is highlighted by increasing evidence on the impacts of recurrent worldwide oil spills in this sensitive species during early development. By means of high throughput sequencing, we characterized the developmental transcriptome of mahi-mahi at three critical developmental stages, from pharyngula embryonic stage (24 hpf) to 48 hpf yolk-sac larva (transition 1), and to 96 hpf free-swimming larva (transition 2). With comparative analysis by multiple bioinformatic tools, a larger number of significantly altered genes and more diverse gene ontology terms were observed during transition 2 than transition 1. Cellular and tissue development terms were more significantly enriched in transition 1, while metabolism related terms were more enriched in transition 2, indicating a switch progressing from general embryonic development to metabolism during the two transitions. Special focus was given on the most significant common canonical pathways (e.g. calcium signaling, glutamate receptor signaling, cAMP response element-binding protein signaling, cardiac β-adrenergic signaling, etc.) and expression of developmental genes (e.g. collagens, myosin, notch, glutamate metabotropic receptor etc.), which were associated with morphological changes of nervous, muscular, and cardiovascular system. These data will provide an important basis for understanding embryonic development and identifying molecular mechanisms of abnormal development in fish species.

Time-dependent changes and potential mechanisms of glucose-lipid metabolic disorders associated with chronic clozapine or olanzapine treatment in rats

Chronic treatment with second-generation antipsychotic drugs (SGAs) has been associated with an increased risk of metabolic syndrome. To evaluate the longitudinal changes in glucose-lipid homeostasis after SGA use, we studied the time-dependent effects of olanzapine (OLZ) (3 mg/kg, b.i.d.) or clozapine (CLZ) (20 mg/kg, b.i.d.) treatment on metabolic profiles for 9 weeks in rats. Although only OLZ significantly increased body weight in rats, both OLZ and CLZ elevated blood lipid levels. Chronic OLZ treatment induced significant weight gain leading to a higher fasting insulin level and impaired glucose tolerance, whereas CLZ lowered fasting insulin levels and impaired glucose tolerance independent of weight gain. Treatment with both drugs deranged AKT/GSK phosphorylation and up-regulated muscarinic M3 receptors in the rats’ livers. Consistent with an elevation in lipid levels, both OLZ and CLZ significantly increased the protein levels of nuclear sterol regulatory element-binding proteins (SREBPs) in the liver, which was associated with improvement in hepatic histamine H1R. However, enhanced carbohydrate response element binding protein (ChREBP) signalling was observed in only CLZ-treated rats. These results suggest that SGA-induced glucose-lipid metabolic disturbances could be independent of weight gain, possibly through activation of SREBP/ChREBP in the liver.

RhoA inhibits the hypoxia-induced apoptosis and mitochondrial dysfunction in chondrocytes via positively regulating the CREB phosphorylation.

Chondrocytes that are embedded within the growth plate or the intervertebral disc are sensitive to environmental stresses, such as inflammation and hypoxia. However, little is known about the molecular signalling pathways underlining the hypoxia-induced mitochondrial dysfunction and apoptosis in chondrocytes. In the present study, we firstly examined the hypoxia-induced apoptosis, mitochondrial dysfunction and the activation of cyclic adenosine monophosphate (cAMP) response element-binding protein (CREB) signalling in human chondrocyte cell line, C28/I2 and then investigated the regulatory role of RhoA, a well-recognized apoptosis suppressor, in such process, with gain-of-function strategy. Our results indicated that hypoxia induced apoptosis and inhibited CREB phosphprylation in chondrocytes, meanwhile, the dysfunctional mitochondria with up-regulated mitochondrial superoxide and reactive oxygen species (ROS) levels, whereas with a reduced mitochondrial membrane potential (MMP) and Complex IV activity were observed in the hypoxia-treated C28/I2 cells. However, the overexpressed RhoA blocked the hypoxia-mediated reduction in CREB phosphprylation and inhibited the apoptosis induction, along with an ameliorated mitochondrial function in the hypoxia-treated C28/I2 cells. In conclusion, the present study confirmed the reduced CREB phosphorylation, along with the apoptosis induction and mitochondrial dysfunction in the hypoxia-treated chondrocyte cells. And the overexpression of RhoA ameliorated the hypoxia-induced mitochondrial dysfunction and apoptosis via blocking the hypoxia-mediated reduction in CREB phosphorylation.

Ginsenoside Rg3 exerts anti-depressive effect on an NMDA-treated cell model and a chronic mild stress animal model

Depression is a common mental disorder and a leading cause of disability. At its most severe, it can lead to suicide. Recently, there has been growing interest in the application of natural herbs for the prevention and treatment of depression. In this report, we found that the ginsenoside active component Rg3 has an apparent antidepressant effect. In N-methyl-d-aspartic acid (NMDA)-treated HT22 murine hippocampal neuronal cells, Rg3 recovered proliferation and inhibited apoptosis by altering the cell cycle. More interestingly, Rg3 led to apparent physiological behavior change in a chronic mild stress model as seen in forced swim, tail suspension, and sucrose preference tests. This effect was mediated by the phosphorylation of cAMP response element binding protein (CREB) and brain-derived neurotrophic factor (BDNF) signaling. This study provides direct evidence to support the antidepressant effects of ginsenoside Rg3, potentially indicating its application in the treatment of clinical depression.

The involvement of BDNF-CREB signaling pathways in the pharmacological mechanism of combined SSRI- antipsychotic treatment in schizophrenia

Previous studies into the mechanism of SSRI-antipsychotic synergism in our laboratory identified unique changes in the brain, particularly in the γ-aminobutyric acid (GABA)-A receptor and its modulators. This study examined the role of brain derived neurotrophic factor (BDNF)-cAMP response element binding (CREB) protein signaling pathways, including protein kinase B (AKT), glycogen synthase kinase (GSK)-3β and related molecules in the molecular response to haloperidol, fluvoxamine, combined haloperidol+fluvoxamine and clozapine treatments in rat frontal cortex, hippocampus and primary cortical neuronal cultures. The effect of fluvoxamine augmentation on BDNF-CREB pathways in peripheral mononuclear cells (PMC׳s) of medicated schizophrenia patients was also studied. Chronic haloperidol (1mg/kg) +fluvoxamine (10mg/kg) treatment increased TrkB receptor and BDNF expression levels, and the phosphorylation of AKT/CREB/GSK-3β, compared to the individual drugs in rat brain. In addition, haloperidol+fluvoxamine treatment improved cognitive functions in rats, indicating that the molecular changes may have a role in behavioral improvement. In primary neuronal cell cultures, pretreatment with a selective PI3K inhibitor abolished the haloperidol+fluvoxamine-induced phosphorylation of AKT and GSK-3β, but did not affect the upregulation of CREB phosphorylation. In the clinic, PMC׳s of treated patients showed upregulation of mRNA expression and protein levels of BDNF, CREB and AKT after addition of fluvoxamine. Analyses of PMC genes and proteins showed significant inter-correlations and some gene changes correlated with improvement in negative and cognitive symptoms. Our study provides new knowledge of the molecular mechanisms of symptom amelioration in schizophrenia and may advance development of new drugs for this disease and other neuropsychiatric disorders.

Maternal low-protein diet decreases brain-derived neurotrophic factor expression in the brains of the neonatal rat offspring

Prenatal exposure to a maternal low-protein (LP) diet has been known to cause cognitive impairment, learning and memory deficits. However, the underlying mechanisms have not been identified. Herein, we demonstrate that a maternal LP diet causes, in the brains of the neonatal rat offspring, an attenuation in the basal expression of the brain-derived neurotrophic factor (BDNF), a neurotrophin indispensable for learning and memory. Female rats were fed either a 20% normal protein (NP) diet or an 8% LP 3 weeks before breeding and during the gestation period. Maternal LP diet caused a significant reduction in the Bdnf expression in the brains of the neonatal rats. We further found that the maternal LP diet reduced the activation of the cAMP/protein kinase A/cAMP response element binding protein (CREB) signaling pathway. This reduction was associated with a significant decrease in CREB binding to the Bdnf promoters. We also show that prenatal exposure to the maternal LP diet results in an inactive or repressed exon I and exon IV promoter of the Bdnf gene in the brain, as evidenced by fluxes in signatory hallmarks in the enrichment of acetylated and trimethylated histones in the nucleosomes that envelop the exon I and exon IV promoters, causing the Bdnf gene to be refractory to transactivation. Our study is the first to determine the impact of a maternal LP diet on the basal expression of BDNF in the brains of the neonatal rats exposed prenatally to an LP diet.

CREB Signaling Is Involved in Rett Syndrome Pathogenesis.

Rett syndrome (RTT) is a debilitating neurodevelopmental disorder caused by mutations in the MECP2 gene. To facilitate the study of cellular mechanisms in human cells, we established several human stem cell lines: human embryonic stem cell (hESC) line carrying the common T158M mutation (MECP2T158M/T158M ), hESC line expressing no MECP2 (MECP2-KO), congenic pair of wild-type and mutant RTT patient-specific induced pluripotent stem cell (iPSC) line carrying the V247fs mutation (V247fs-WT and V247fs-MT), and iPSC line in which the V247fs mutation was corrected by CRISPR/Cas9-based genome editing (V247fs-MT-correction). Detailed analyses of forebrain neurons differentiated from these human stem cell lines revealed genotype-dependent quantitative phenotypes in neurite growth, dendritic complexity, and mitochondrial function. At the molecular level, we found a significant reduction in the level of CREB and phosphorylated CREB in forebrain neurons differentiated from MECP2T158M/T158M , MECP2-KO, and V247fs-MT stem cell lines. Importantly, overexpression of CREB or pharmacological activation of CREB signaling in those forebrain neurons rescued the phenotypes in neurite growth, dendritic complexity, and mitochondrial function. Finally, pharmacological activation of CREB in the female Mecp2 heterozygous mice rescued several behavioral defects. Together, our study establishes a robust in vitro platform for consistent quantitative evaluation of genotype-dependent RTT phenotypes, reveals a previously unappreciated role of CREB signaling in RTT pathogenesis, and identifies a potential therapeutic target for RTT.SIGNIFICANCE STATEMENT Our study establishes a robust human stem cell-based platform for consistent quantitative evaluation of genotype-dependent Rett syndrome (RTT) phenotypes at the cellular level. By providing the first evidence that enhancing cAMP response element binding protein signaling can alleviate RTT phenotypes both in vitro and in vivo, we reveal a previously unappreciated role of cAMP response element binding protein signaling in RTT pathogenesis, and identify a potential therapeutic target for RTT.

Effect of sevoflurane on hippocampal cAMP-PKA-CREB signaling pathway in aged rats

Objective To evaluate the effect of sevoflurane on hippocampal cyclic AMP (cAMP)-protein kinase A (PKA)-cAMP response element-binding protein (CREB) signaling pathway in aged rats. Methods Sixty healthy aged male Sprague-Dawley rats, aged 18 months, weighing 600-750 g, were divided into control group (C group, n=30) and sevoflurane group (group Sev, n=30) using a random number table method.Group Sev inhaled 2% sevoflurane for 4 h. Group C inhaled the mixture of 50% air and oxygen (2 L/min) for 4 h. Morris water maze test was performed on 1-6 days before anesthesia and at 1 day after the end of anesthesia (at the corresponding time points in group C). The animals were sacrificed and hippocampi were removed at 1, 3 and 7 days after anesthesia (at the corresponding time points in group C) for determination of the hippocampal cAMP content (using enzyme-linked immunosorbent assay) and expression of hippocampal CREB, phosphorylated CREB (p-CREB) and PKA (using Western blot), and the p-CREB/CREB ratio was calculated. Results Compared with group C, the escape latency and swimming distance were significantly prolonged, the frequency of crossing the original platform was decreased, the time of staying at the original platform quadrant was shortened at 1 day after the end of anesthesia, and the expression of hippocampal cAMP and PKA was down-regulated at 1, 3 and 7 days after the end of anesthesia, and the expression of CREB and p-CREB was down-regulated and p-CREB/CREB ratio was decreased at 1 day after the end of anesthesia in group Sev (P<0.05). Conclusion The mechanism of sevoflurane-induced postoperative cognitive dysfunction may be related to inhibiting hippocampal cAMP-PKA-CREB signaling pathway in aged rats. Key words: Cyclic AMP; Cyclic AMP-dependent protein kinases; Cyclic AMP response element-binding protein; Anesthetics, inhalation; Cognition disorders

IP-10/CXCR3 Axis Promotes the Proliferation of Vascular Smooth Muscle Cells through ERK1/2/CREB Signaling Pathway

Excessive proliferation of vascular smooth muscle cells is one of the main pathological processes leading to atherosclerosis and intimal hyperplasia after vascular interventional therapy. Our previous study has shown that interferon-γ inducible protein-10 contributes to the proliferation of vascular smooth muscle cell. However, the underlying mechanisms remain unclear. Extracellular signal-regulated kinase 1/2, serine/threonine kinase Akt, and cAMP response element binding protein are signaling pathways, which are considered to play important roles in the processes of vascular smooth muscle cell proliferation. Moreover, chemokine receptor 3 and Toll-like receptor 4 are potential receptors of inducible protein-10 in this process. In the present study, IP-10 was found to directly induce vascular smooth muscle cell proliferation, and exposure to inducible protein-10 activated extracellular signal-regulated kinase 1/2, serine/threonine kinase, and cAMP response element binding protein signaling. Inhibitor of extracellular signal-regulated kinase 1/2, rather than inhibitor of serine/threonine kinase, inhibited the phosphorylation of cAMP response element binding protein and reduced inducible protein-10-stimulated vascular smooth muscle cell proliferation. Knockdown of cAMP response element binding protein by siRNA inhibited inducible protein-10-induced vascular smooth muscle cell proliferation. Moreover, anti-CXCR3 IgG, instead of anti-Toll-like receptor 4 IgG, reduced inducible protein-10-induced vascular smooth muscle cell proliferation and inducible protein-10-stimulated extracellular signal-regulated kinase 1/2 and cAMP response element binding protein activation. Together, these results indicate that inducible protein-10 promotes vascular smooth muscle cell proliferation via chemokine receptor 3 and activation of extracellular signal-regulated kinase 1/2 inducible protein-10-induced vascular smooth muscle cell proliferation. These data provide important targets for future studies to modulate atherosclerosis and restenosis after vascular interventional therapy.

Mechanisms Underlying the Antidepressant Response of Acupuncture via PKA/CREB Signaling Pathway.

Protein kinase A (PKA)/cAMP response element-binding (CREB) protein signaling pathway, contributing to impaired neurogenesis parallel to depressive-like behaviors, has been identified as the crucial factor involved in the antidepressant response of acupuncture. However, the molecular mechanisms associated with antidepressant response of acupuncture, neurogenesis, and depressive-like behaviors ameliorating remain unexplored. The objective was to identify the mechanisms underlying the antidepressant response of acupuncture through PKA signaling pathway in depression rats by employing the PKA signaling pathway inhibitor H89 in in vivo experiments. Our results indicated that the expression of hippocampal PKA-α and p-CREB was significantly downregulated by chronic unpredicted mild stress (CUMS) procedures. Importantly, acupuncture reversed the downregulation of PKA-α and p-CREB. The expression of PKA-α was upregulated by fluoxetine, but not p-CREB. No significant difference was found between Acu and FLX groups on the expression of PKA-α and p-CREB. Interestingly, H89 inhibited the effects of acupuncture or fluoxetine on upregulating the expression of p-CREB, but not PKA-α. There was no significant difference in expression of CREB among the groups. Conclusively, our findings further support the hypothesis that acupuncture could ameliorate depressive-like behaviors by regulating PKA/CREB signaling pathway, which might be mainly mediated by regulating the phosphorylation level of CREB.

Roscovitine, a CDK5 Inhibitor, Alleviates Sevoflurane-Induced Cognitive Dysfunction via Regulation Tau/GSK3β and ERK/PPARγ/CREB Signaling

Background/Aims: Multiple exposures to anesthesia in children may increase the risk of developing cognitive impairment. Sevoflurane is an anesthetic that is commonly used in children during surgery. Cyclin-dependent kinase (CDK) 5 is involved in the regulation of sevoflurane-induced cognitive dysfunction, but the mechanistic details remain unclear. The present study evaluated the mechanism by which CDK5 mediates sevoflurane-induced cognitive dysfunction in mice. Methods: Hippocampal neurons were isolated from postnatal day 0 C57BL/6 mouse pups. Six-day-old wild-type mice were exposed to sevoflurane and then treated with the CDK5 inhibitor roscovitine. The effects on cognitive function were evaluated with the Morris water maze and neuronal damage in the hippocampus was assessed by immunohistochemical analysis. Results: CDK5 activation increased neuronal damage by inducing Tau/glycogen synthase kinase (GSK) 3β and suppressing extracellular signal-regulated kinase (ERK)/peroxisome proliferator-activated receptor (PPAR)γ/cyclic AMP response element-binding protein (CREB) signaling following exposure to sevoflurane. CDK5 inhibition by roscovitine administration alleviated sevoflurane-induced neuronal damage and cognitive impairment. Conclusions: Inhibiting CDK5 with roscovitine has neuroprotective effects against neuronal injury and cognitive dysfunction caused by sevoflurane anesthesia that are exerted via modulation of Tau/GSK3β and ERK/PPARγ/CREB signaling.

Heat induces interleukin-6 in skeletal muscle cells via TRPV1/PKC/CREB pathways.

Interleukin-6 (IL-6) is released from skeletal muscle cells and induced by exercise, heat, catecholamine, glucose, lipopolysaccharide, reactive oxygen species, and inflammation. However, the mechanism that induces release of IL-6 from skeletal muscle cells remains unknown. Thermosensitive transient receptor potential (TRP) proteins such as TRPV1-4 play vital roles in cellular functions. In this study we hypothesized that TRPV1 senses heat, transmits a signal into the nucleus, and produces IL-6. The purpose of the present study is to investigate the underlying mechanisms whereby skeletal muscle cells sense and respond to heat. When mouse myoblast cells were exposed to 37-42°C for 2 h, mRNA expression of IL-6 increased in a temperature-dependent manner. Heat also increased IL-6 secretion in myoblast cells. A fura 2 fluorescence dual-wavelength excitation method showed that heat increased intracellular calcium flux in a temperature-dependent manner. Intracellular calcium flux and IL-6 mRNA expression were increased by the TRPV1 agonists capsaicin and N-arachidonoyldopamine and decreased by the TRPV1 antagonists AMG9810 and SB366791 and siRNA-mediated knockdown of TRPV1. TRPV2, 3, and 4 agonists did not change intracellular calcium flux. Western blotting with inhibitors demonstrated that heat increased phosphorylation levels of TRPV1, followed by PKC and cAMP response element-binding protein (CREB). PKC inhibitors, Gö6983 and staurosporine, CREB inhibitors, curcumin and naphthol AS-E, and knockdown of CREB suppressed the heat-induced increases in IL-6. These results indicate that heat increases IL-6 in skeletal muscle cells through the TRPV1, PKC, and CREB signal transduction pathway.NEW & NOTEWORTHY Heat increases the release of interleukin-6 (IL-6) from skeletal muscle cells. IL-6 has been shown to serve immune responses and metabolic functions in muscle. It can be anti-inflammatory as well as proinflammatory. However, the mechanism that induces release of IL-6 from skeletal muscle cells remains unknown. Here we show that heat increases IL-6 in skeletal muscle cells through the transient receptor potential vannilloid 1, PKC, and cAMP response element-binding protein signal transduction pathway.

Cigarette smoke extracts induce overexpression of the proto-oncogenic gene interleukin-13 receptor α2 through activation of the PKA-CREB signaling pathway to trigger malignant transformation of lung vascular endothelial cells and angiogenesis

Cigarette smoking is a major cause of lung cancer. Tumor-associated endothelial cells (TAECs) play important roles in tumor angiogenesis and metastasis. However, whether cigarette smoking can trigger genesis of lung TAECs has not been reported yet. In the current study, we used lung endothelial cell (EC) lines as a model to study the pathological effect of cigarette smoke extracts (CSEs) on human lung ECs, and found that a lower dose of 4% CSEs obviously caused abnormal morphological changes in ECs, increased the permeability of endothelial monolayer, while a higher concentration of 8% CSEs caused EC apoptosis. Strikingly, CSEs induced a 117-fold overexpression of a pro-tumorigenic interleukin-13 receptor α2 gene (IL-13Rα2, also named as CT-19) through activation of the protein kinase A (PKA) and cAMP response element-binding protein (CREB) signaling pathway. A PKA specific inhibitor H89 completely abolished CSEs-induced IL-13Rα2 overexpression. The overexpression of IL-13Rα2 in lung ECs significantly increased the tumorigenic, migratory, and angiogenic capabilities of the cells, suggesting that IL-13Rα2 promotes genesis of lung TAECs. Together, our data show that CSEs activate the PKA, CREB, and IL-13Rα2 axis in lung ECs, and IL-13Rα2 promotes the malignant transformation of lung ECs and genesis of TAECs with robust angiogenic and oncogenic capabilities. Our study provides new insight into the mechanism of CSEs-triggered lung cancer angiogenesis and tumorigenesis, suggesting that the PKA-CREB-IL-13Rα2 axis is a potential target for novel anti-lung tumor angiogenesis and anti-lung cancer drug discovery.

Adaptive Activation of a Stress Response Pathway Improves Learning and Memory Through Gs and β-Arrestin-1–Regulated Lactate Metabolism

BackgroundStress is a conserved physiological response in mammals. Whereas moderate stress strengthens memory to improve reactions to previously experienced difficult situations, too much stress is harmful. MethodsWe used specific β-adrenergic agonists, as well as β2-adrenergic receptor (β2AR) and arrestin knockout models, to study the effects of adaptive β2AR activation on cognitive function using Morris water maze and object recognition experiments. We used molecular and cell biological approaches to elucidate the signaling subnetworks. ResultsWe observed that the duration of the adaptive β2AR activation determines its consequences on learning and memory. Short-term formoterol treatment, for 3 to 5 days, improved cognitive function; however, prolonged β2AR activation, for more than 6 days, produced harmful effects. We identified the activation of several signaling networks downstream of β2AR, as well as an essential role for arrestin and lactate metabolism in promoting cognitive ability. Whereas Gs–protein kinase A–cyclic adenosine monophosphate response element binding protein signaling modulated monocarboxylate transporter 1 expression, β-arrestin-1 controlled expression levels of monocarboxylate transporter 4 and lactate dehydrogenase A through the formation of a β-arrestin-1/phospho-mitogen-activated protein kinase/hypoxia-inducible factor-1α ternary complex to upregulate lactate metabolism in astrocyte-derived U251 cells. Conversely, long-term treatment with formoterol led to the desensitization of β2ARs, which was responsible for its decreased beneficial effects. ConclusionsOur results not only revealed that β-arrestin-1 regulated lactate metabolism to contribute to β2AR functions in improved memory formation, but also indicated that the appropriate management of one specific stress pathway, such as through the clinical drug formoterol, may exert beneficial effects on cognitive abilities.

The Schizophrenia-Associated BRD1 Gene Regulates Behavior, Neurotransmission, and Expression of Schizophrenia Risk Enriched Gene Sets in Mice

BackgroundThe schizophrenia-associated BRD1 gene encodes a transcriptional regulator whose comprehensive chromatin interactome is enriched with schizophrenia risk genes. However, the biology underlying the disease association of BRD1 remains speculative. MethodsThis study assessed the transcriptional drive of a schizophrenia-associated BRD1 risk variant in vitro. Accordingly, to examine the effects of reduced Brd1 expression, we generated a genetically modified Brd1+/− mouse and subjected it to behavioral, electrophysiological, molecular, and integrative genomic analyses with focus on schizophrenia-relevant parameters. ResultsBrd1+/− mice displayed cerebral histone H3K14 hypoacetylation and a broad range of behavioral changes with translational relevance to schizophrenia. These behaviors were accompanied by striatal dopamine/serotonin abnormalities and cortical excitation-inhibition imbalances involving loss of parvalbumin immunoreactive interneurons. RNA-sequencing analyses of cortical and striatal micropunches from Brd1+/− and wild-type mice revealed differential expression of genes enriched for schizophrenia risk, including several schizophrenia genome-wide association study risk genes (e.g., calcium channel subunits [Cacna1c and Cacnb2], cholinergic muscarinic receptor 4 [Chrm4)], dopamine receptor D2 [Drd2], and transcription factor 4 [Tcf4]). Integrative analyses further found differentially expressed genes to cluster in functional networks and canonical pathways associated with mental illness and molecular signaling processes (e.g., glutamatergic, monoaminergic, calcium, cyclic adenosine monophosphate [cAMP], dopamine- and cAMP-regulated neuronal phosphoprotein 32 kDa [DARPP-32], and cAMP responsive element binding protein signaling [CREB]). ConclusionsOur study bridges the gap between genetic association and pathogenic effects and yields novel insights into the unfolding molecular changes in the brain of a new schizophrenia model that incorporates genetic risk at three levels: allelic, chromatin interactomic, and brain transcriptomic.

GIV/Girdin activates Gαi and inhibits Gαs via the same motif

We previously showed that guanine nucleotide-binding (G) protein α subunit (Gα)-interacting vesicle-associated protein (GIV), a guanine-nucleotide exchange factor (GEF), transactivates Gα activity-inhibiting polypeptide 1 (Gαi) proteins in response to growth factors, such as EGF, using a short C-terminal motif. Subsequent work demonstrated that GIV also binds Gαs and that inactive Gαs promotes maturation of endosomes and shuts down mitogenic MAPK-ERK1/2 signals from endosomes. However, the mechanism and consequences of dual coupling of GIV to two G proteins, Gαi and Gαs, remained unknown. Here we report that GIV is a bifunctional modulator of G proteins; it serves as a guanine nucleotide dissociation inhibitor (GDI) for Gαs using the same motif that allows it to serve as a GEF for Gαi. Upon EGF stimulation, GIV modulates Gαi and Gαs sequentially: first, a key phosphomodification favors the assembly of GIV-Gαi complexes and activates GIV's GEF function; then a second phosphomodification terminates GIV's GEF function, triggers the assembly of GIV-Gαs complexes, and activates GIV's GDI function. By comparing WT and GIV mutants, we demonstrate that GIV inhibits Gαs activity in cells responding to EGF. Consequently, the cAMP→PKA→cAMP response element-binding protein signaling axis is inhibited, the transit time of EGF receptor through early endosomes are accelerated, mitogenic MAPK-ERK1/2 signals are rapidly terminated, and proliferation is suppressed. These insights define a paradigm in G-protein signaling in which a pleiotropically acting modulator uses the same motif both to activate and to inhibit G proteins. Our findings also illuminate how such modulation of two opposing Gα proteins integrates downstream signals and cellular responses.

Effect of Kaempferia parviflora extract and its polymethoxyflavonoid components on testosterone production in mouse testis-derived tumour cells

Low testosterone is associated with late-onset hypogonadism, age-related disease, type 2 diabetes, and cardiovascular disease. In this study, we investigated the effects of Kaempferia parviflora extract and two of its components, 5,7-dimethoxyflavone and 5-hydroxy-3,7,3′,4′-tetramethoxyflavone, on testosterone production in mouse testis-derived tumour cells I-10. K. parviflora fractions containing 5,7-dimethoxyflavone elicited testosterone production in these cells, as did nobiletin, a polymethoxyflavonoid found in citrus species, but not K. parviflora, which was used as a comparative control. Furthermore, we found that these compounds enhanced testosterone production via cyclic AMP (cAMP)/cAMP response element binding protein signalling. In particular, the compounds inhibited activation of phosphodiesterase and thereby increased production of cAMP. These findings support the possibility that K. parviflora extract, 5,7-dimethoxyflavone, and nobiletin may mitigate age-related disease and hypogonadism by enhancing testosterone production.

Reduced pCREB in Alzheimer's disease prefrontal cortex is reflected in peripheral blood mononuclear cells.

Cyclic-AMP response element-binding protein (CREB) signaling has a critical role in the formation of memories. CREB signaling is dysfunctional in the brains of mouse models of Alzheimer's disease (AD), and evidence suggests that CREB signaling may be disrupted in human AD brains as well. Here, we show that both CREB and its activated form pCREB-Ser133 (pCREB) are reduced in the prefrontal cortex of AD patients. Similarly, the transcription cofactors CREB-binding protein (CBP) and p300 are reduced in the prefrontal cortex of AD patients, indicating additional dysfunction of CREB signaling in AD. Importantly, we show that pCREB expression is reduced in peripheral blood mononuclear cells (PBMC) of AD subjects. In addition, pCREB levels in PBMC positively correlated with pCREB expression in the postmortem brain of persons with AD. These results suggest that pCREB expression in PBMC may be indicative of its expression in the brain, and thus offers the intriguing possibility of pCREB as a biomarker of cognitive function and disease progression in AD.

Mechanical Strain Regulates Osteoblast Proliferation Through Ca2+-CaMK-CREB Signal Pathway

Objective To investigate the effects of mechanical strain on Ca2+-calmodulin dependent kinase (CaMK)-cAMP response element binding protein (CREB) signal pathway and proliferation of osteoblasts.Methods Using a four-point bending device, MC3T3-E1 cells were exposed to mechanical tensile strains of 2500 µs and 5000 µs at 0.5 Hz respectively. The intracellular free Ca2+ ([Ca2+]i) concentration and calmodulin activity were assayed by fluorospectrophotometry, CaMK II β, CREB, and phosphorylated (activated) CREB (p-CREB) were assessed by Western blot, and cells proliferation was assayed with MTT. Pretreatment with verapamil was carried out to block Ca2+ channel, and inhibitor U73122 was used to inhibit phospholipase C (PLC).Results Mechanical strains of 2500 µs and 5000 µs for 1 to 10 minutes both increased [Ca2+]i level of the cells. The 2500 µs strain, a periodicity of 1 h/d for 3 days, activated calmodulin, elevated protein levels of CaMK II β and p-CREB, and promoted cells proliferation, which were attenuated by pretreatment of verapamil or U73122. The effects of 5000 µs strain on calmodulin, CaMK II β, p-CREB and proliferation were contrary to 2500 µs strain.Conclusion The mechanical strain regulates osteoblasts proliferation through Ca2+-CaMK-CREB signal pathway via Ca2+ channel and PLC/IP3 transduction cascades.

Wnt3a regulates mitochondrial biogenesis through p38/CREB pathway

Wnt3a is established as an important regulator of various developmental processes, especially in osteogenesis, adipogenesis and mitochondrial biogenesis. Numerous studies reported Wnt3a regulates osteogenesis and adipogenesis, but the mechanisms by which Wnt3a regulates mitochondrial biogenesis are not well understood. In this study, results suggested that Wnt3a stimulates mitochondrial biogenesis by increasing the expression of mitochondrial biogenesis genes and regulators, as well as mitochondrial copy number in adipocytes. As a key mediator of canonical Wnt/β-catenin pathway, β-catenin knockdown had no effect on basal or Wnt3a-mediated mitochondrial biogenesis in adipocytes, which suggested that Wnt3a-mediated mitochondrial biogenesis was independent of β-catenin-dependent canonical Wnt/β-catenin pathway. However, Wnt3a inhibited p38/CREB (p38 mitogen-activated protein kinase/cAMP response element-binding protein) signaling activation and p38 inhibitor impaired Wnt3a-stimulated mitochondrial biogenesis, indicating p38/CREB pathway could be involved in the regulation of Wnt3a-mediated mitochondrial biogenesis in adipocytes. In conclusion, our data showed that Wnt3a stimulates mitochondrial biogenesis in adipocytes, which is at least partially through activation of p38/CREB pathway.