Tachypacing-induced CREB/CD44 signaling contributes to the suppression of L-type calcium channel expression and the development of atrial remodeling.

BACKGROUND: Atrial fibrillation (AF), a common arrhythmia in clinics, is characterized as downregulation of L-type calcium channel (LTCC) and shortening of atrial action potential duration (APD). Our prior studies have shown the association of CD44 with AF genesis. OBJECTIVE: The purpose of this study was to explore the potential role of CD44 and its related signaling in tachypacing-induced downregulation of LTCC. METHODS AND RESULTS: In vitro, tachypacing in atrium-derived myocytes (HL-1 cell line) induced activation (phosphorylation) of cyclic adenosine monophosphate response element-binding protein (CREB). Furthermore, tachypacing promoted an association between CREB and CD44 in HL-1 myocytes, which was documented in atrial tissues from patients with AF. Deletion and mutational analysis of the LTCC promoter along with chromatin immunoprecipitation revealed that cyclic adenosine monophosphate response element is essential for tachypacing-inhibited LTCC transcription. Tachypacing also hindered the binding of p-CREB to the promoter of LTCC. Blockade of CREB/CD44 signaling in HL-1 cells attenuated tachypacing-triggered downregulation of LTCC and shortening of APD. Atrial myocytes isolated from CD44-/- mice exhibited higher LTCC current and longer APD than did those from wild-type mice. Ex vivo, tachypacing caused less activation of CREB in CD44-/- mice than in wild-type mice. In vivo, burst atrial pacing stimulated less inducibility of AF in CREB inhibitor-treated mice than in controls. CONCLUSION: Tachypacing-induced CREB/CD44 signaling contributes to the suppression of LTCC, which provides valuable information about the pathogenesis of atrial modeling and AF.

Gene expression meta-analysis reveals the up-regulation of CREB1 and CREBBP in Brodmann Area 10 of patients with schizophrenia.

Cognitive impairments characterize individuals with schizophrenia, and are correlated to the patients' functional outcome. The transcription factor Cyclic AMP-responsive element-binding protein-1 (CREB1) is involved in learning and memory processes. CREB1 and both CREB-binding protein (CREBBP) and E1A Binding Protein P300 (EP300), co-activators of CREB1, have been associated with schizophrenia. We performed a systematic meta-analysis of CREB1, CREBBP and EP300 differential expression in post mortem Brodmann Area 10 (BA10) samples of patients with schizophrenia vs. healthy controls, following the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines. Two microarray datasets met the inclusion criteria (overall 41 schizophrenia samples and 38 controls were analyzed). We detect up-regulation of CREB1 and CREBBP in BA10 samples of patients with schizophrenia, while EP300 wasn't differentially expressed. The integration of two independent datasets and the positive correlation between the expression patterns of CREB1 and CREBBP increase the validity of the results. The up-regulation of CREB1 and its co-activator CREBBP might relate to BA10 altered activation that has been shown in schizophrenia. As BA10 was shown to be involved in the cognitive impairments associated with schizophrenia, this suggests involvement of CREB1 and CREBBP in the cognitive symptoms that characterize the disease.

Genetic polymorphism and transcriptional regulation of CREBBP gene in patient with diffuse large B-cell lymphoma.

In the present study, we aim to examine the relationship between genetic polymorphism and transcriptional expression of cyclic AMP response element binding protein (CREBBP) and the risk of diffuse large B-cell lymphoma (DLBCL). Two hundred and fifty healthy individuals and 248 DLBCL patients participated in the present study. The CREBBP rs3025684 polymorphism was detected by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). The mRNA expression of CREBBP was tested by the real-time quantitative PCR (RT-qPCR). The allele A frequency of CREBBP rs3025684 in DLBCL patients was obviously higher than that of controls (P=0.01). No significant difference was detected between CREBBP rs3025684 polymorphism and clinical characteristics of DLBCL patients when subgrouped according to different parameters. The results demonstrated that the allele A of CREBBP rs3025684 increased the susceptibility to DLBCL (P=0.004), with a worse overall survival (OS) rate (P=0.002), a worse progression-free survival (PFS) rate (P=0.033) and poor prognosis (P=0.003) in DLCBL patients. Furthermore, the expression of CREBBP mRNA was considerably decreased in DLBCL patients as compared with controls (P<0.001), and the expression in patients with GG genotype was up-regulated in comparison with patients with GA and AA genotype (P=0.016 and P=0.001, respectively). However, no statistical differences were found in OS (P=0.201) and PFS (P=0.353) between the lower CREBBP mRNA level subgroup and higher CREBBP mRNA level subgroup. These data suggested that the CREBBP gene may be an important prognostic factor in DLBCL patients and perform an essential function in the development of DLBCL.

Expression of Hypoxia Inducible Factor 1alpha Is Protein Kinase A-dependent in Primary Cortical Astrocytes Exposed to Severe Hypoxia.

The hypoxia inducible factor 1 (HIF-1) and the cyclic AMP-responsive element binding protein (CREB) are two transcription factors that have been studied in the context of neuronal survival and neurodegeneration. HIF-1 upregulation and CREB activation have been observed not only in neurons but also in astrocytes under conditions of hypoxia. We hypothesized that activation of CREB regulate HIF-1α expression in the nucleus of cortical astrocytes under in vitro ischemic condition. To test the hypothesis, we determined the effects of inhibiting the CREB activation pathway on the expression of HIF-1α protein in astrocytes exposed to CoCl2 and severe hypoxia (near anoxia, 0.1% O2). The results demonstrated that inhibition of CaMKII and CaMKIV had no effect on both HIF-1α and pCREB expression in cortical astrocytes exposed to CoCl2 and anoxia. In contrast, PKA inhibition lowered the expression of HIF-1α and pCREB expression. Furthermore, the inhibition of PKA but not CaMKII or CaMKIV increased cell death of astrocytes exposed to near anoxia. The results suggest that PKA plays an important role in the cell survival signaling pathways in astrocytes.

Estradiol up-regulates L-type Ca2+ channels via membrane-bound estrogen receptor/phosphoinositide-3-kinase/Akt/cAMP response element-binding protein signaling pathway.

BACKGROUND: In long QT syndrome type 2, women are more prone than men to the lethal arrhythmia torsades de pointes. We previously reported that 17ß-estradiol (E2) up-regulates L-type Ca2+ channels and current (ICa,L) (∼30%) in rabbit ventricular myocytes by a classic genomic mechanism mediated by estrogen receptor-α (ERα). In long QT syndrome type 2 (IKr blockade or bradycardia), the higher Ca2+ influx via ICa,L causes Ca2+ overload, spontaneous sarcoplasmic reticulum Ca2+ release, and reactivation of ICa,L that triggers early afterdepolarizations and torsades de pointes. OBJECTIVE: The purpose of this study was to investigate the molecular mechanisms whereby E2 up-regulates ICa,L, which are poorly understood. METHODS: H9C2 and rat myocytes were incubated with E2 ± ER antagonist, or inhibitors of downstream transcription factors, for 24 hours, followed by western blots of Cav1.2α1C and voltage-clamp measurements of ICa,L. RESULTS: Incubation of H9C2 cells with E2 (10-100 nM) increased ICa,L density and Cav1.2α1C expression, which were suppressed by the ER antagonist ICI182,780 (1 µM). Enhanced ICa,L and Cav1.2α1C expression by E2 was suppressed by inhibitors of phosphoinositide-3-kinase (Pi3K) (30 µM LY294002; P <.05) and Akt (5 µM MK2206) but not of mitogen-activated protein kinase (5 µM U0126) or protein kinase A (1 µM KT5720). E2 incubation increased p-CREB via the Pi3K/Akt pathway, reached a peak in 20 minutes (3-fold), and leveled off to 1.5-fold 24 hours later. Furthermore, a CREB decoy oligonucleotide inhibited E2-induced Cav1.2α1C expression, whereas membrane-impermeable E2 (E2-bovine serum albumin) was equally effective at Cav1.2α1C up-regulation as E2. CONCLUSION: Estradiol up-regulates Cav1.2α1C and ICa,L via plasma membrane ER and by activating Pi3K, Akt, and CREB signaling. The promoter regions of the CACNA1C gene (human-rabbit-rat) contain adjacent/overlapping binding sites for p-CREB and ERα, which suggests a synergistic regulation by these pathways.

An inhibitor of the acetyltransferases CBP/p300 exerts antineoplastic effects on gastrointestinal stromal tumor cells.

Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal neoplasm featured by activated mutations of KIT and PDGFRA. Although overall survival rates have greatly improved by the development of receptor tyrosine kinase inhibitors, most patients ultimately acquire resistance due to secondary mutations of KIT or PDGFRA. Inhibition of the histone acetyltransferases (HATs) CREB­binding protein (CBP) and p300 results in antineoplastic effects in various cancers. To determine whether CBP/p300 can serve as an antineoplastic target for GISTs, specific short interfering RNA sequences and the selective HAT inhibitor C646 were administered to GIST882 cells. Cell viability, apoptosis and the cell cycle were analysed using the Cell Counting Kit-8, a caspase-3/7 activity assay or Annexin V-fluorescein isothiocyanate/propidium iodide (PI) staining and PI staining. Gene and protein expression levels were measured by quantitative real-time polymerase chain reaction and western blotting, respectively. Transcriptional blockage of CBP, rather than p300, resulted in suppression of cell proliferation. Interestingly, both CBP and p300 depletion enhanced caspase-3/7 activity. A lack of CBP and p300 caused ETS translocation variant 1 (ETV1) downregulation and KIT inhibition in GIST cells. Nevertheless, the absence of CBP, not p300, leads to extracellular signal-regulated kinase 1/2 inactivation and c-Jun NH2-terminal kinase activation, suggesting a more crucial role for CBP than p300 in cell proliferation and survival. Furthermore, proliferation of GIST cells was reduced by administration of C646, a selective HAT inhibitor for CBP/p300. Apoptosis induction and cell cycle arrest were detected after exposure to C646, indicating that its antitumor activities were supported by its antiproliferative and proapoptotic effects. Additionally, C646 treatment attenuated ETV1 protein expression and inactivated KIT-dependent pathways. Taken together, the present study suggests that CBP/p300 may serve as novel antineoplastic targets and that use of the selective HAT inhibitor C646 is a promising antitumor strategy for GISTs.

Lactobacillus pentosus var. plantarum C29 ameliorates age-dependent memory impairment in Fischer 344 rats.

UNLABELLED: To understand the anti-inflammaging effect of lactic acid bacteria, we selected NF-κB activation-inhibitory Lactobacillus pentosus var. plantarum C29 and investigated its memory-enhancing and anti-inflammatory effects in aged Fischer 344 rats. C29 (2 × 10(9) CFU rat(-1) ), which was orally administered once a day (6 days per week) for 8 weeks, significantly restored age-reduced spontaneous alternation to 95.2% of that seen in young rats (P < 0.05). C29 treatment also shortened the escape latency on the 4th day to 53.8% of that seen in young rats (P < 0.05). Twenty hours after the last training session, C29 significantly increased the swimming time within the platform quadrant, which was shortened in the aged control rats. Oral administration of C29 restored age-reduced doublecortin (DCX) and brain-derived neurotrophic factor (BDNF) expression and cAMP response element binding protein (CREB) activation in aged rats. Treatment of aged rats with C29 suppressed the expression of p16, cyclooxygenase-2, and inducible nitric oxide synthase, as well as the activation of Akt, mTOR, and NF-κB in the hippocampus. These findings suggest that C29 ameliorates ageing-dependent memory impairment by inhibiting NF-κB signalling pathway, inducing DCX and BDNF expression and activating CREB. SIGNIFICANCE AND IMPACT OF THE STUDY: The anti-inflammatory Lactobacillus pentosus var. plantarum C29 had the memory-enhancing effect in aged Fischer 344 rats by restoring doublecortin and brain-derived neurotrophic factor expression and suppressing p16 expression and NF-κB activation in the brain. Therefore, C29 may be useful in ameliorating age-related degenerative dementia.

Transcriptional coactivator CBP upregulates hTERT expression and tumor growth and predicts poor prognosis in human lung cancers.

Upregulated expression and activation of human telomerase reverse transcriptase (hTERT) is a hallmarker of lung tumorigenesis. However, the mechanism underlying the aberrant hTERT activity in lung cancer cells remains poorly understood. In this study, we found the transcriptional co-activator CBP as a new hTERT promoter-binding protein that regulated hTERT expression and tumor growth in lung adenocarcinoma cells using a biotin-streptavidin-bead pulldown technique. Chromatin immunoprecipitation assay verified the immortalized cell and tumor cell-specific binding of CBP on hTERT promoter. Overexpression of exogenous CBP upregulated the expression of the hTERT promoter-driven luciferase and endogenous hTERT protein in lung cancer cells. Conversely, inhibition of CBP by CBP-specific siRNA or its chemical inhibitor repressed the expression of hTERT promoter-driven luciferase and endogenous hTERT protein as well as telomerase activity. Moreover, inhibition of CBP expression or activity also significantly reduced the proliferation of lung cancer cells in vitro and tumor growth in an xenograft mouse model in vivo. Immunohistochemical analysis of tissue microarrays of lung cancers revealed a positive correlation between CBP and hTERT. Importantly, the patients with high CBP and hTERT expression had a significantly shorter overall survival. Furthermore, CBP was found to interact with and acetylate transactivator Sp1 in lung cancer cells. Inhibition of CBP by CBP-specific siRNA or its chemical inhibitor significantly inhibited Sp1 acetylation and its binding to the hTERT promoter. Collectively, our results indicate that CBP contributes to the upregulation of hTERT expression and tumor growth, and overexpression of CBP predicts poor prognosis in human lung cancers.

A specific CBP/p300-dependent gene expression programme drives the metabolic remodelling in late stages of spermatogenesis.

Histone hyperacetylation is thought to drive the replacement of histones by transition proteins that occur in elongating spermatids (ElS) after a general shut down of transcription. The molecular machineries underlying this histone hyperacetylation remain still undefined. Here, we focused our attention on the role of Cbp and p300 in histone hyperacetylation and in the preceding late-gene transcriptional activity in ElS. A strategy was designed to partially deplete Cbp and p300 in ElS. These cells progressed normally through spermiogenesis and showed normal histone hyperacetylation and removal. However, a genome-wide transcriptomic analysis, performed in the round spermatids (RS) and ElS, revealed the existence of a gene regulatory circuit encompassing genes presenting high expression levels in pre-meiotic cells, undergoing a repressed state in spermatocytes and early post-meiotic cells, but becoming reactivated in ElS, just prior to the global shutdown of transcription. Interestingly, this group of genes was over-represented within the genes affected by Cbp/p300 knock down and were all involved in metabolic remodelling. This study revealed the occurrence of a tightly regulated Cbp/p300-dependent gene expression programme that drives a specific metabolic state both in progenitor spermatogenic cells and in late transcriptionally active spermatids and confirmed a special link between Cpb/p300 and cell metabolism programming previously shown in somatic cells.

Effects of carvedilol treatment on cardiac cAMP response element binding protein expression and phosphorylation in acute coxsackievirus B3-induced myocarditis.

BACKGROUND: The role of ß-adrenergic stimulation on viral myocarditis has been investigated in animal models of viral myocarditis. Excess stimulation of ß-adrenergic receptors by catecholamines causes phosphorylation/activation of cAMP response element binding protein (CREB) by the cAMP signaling pathway. CREB as an important regulator of gene expression mediates the cardiovascular remodeling process and promotes anti-inflammatory immune responses. However, the CREB expression and phosphorylation have not been studied, and the effects of carvedilol (a nonselective ß-adrenoceptor antagonist) on the CREB has not been investigated in the setting of acute viral myocarditis. METHODS: This study was therefore designed to examine the effects of carvedilol on the transcriptional factor CREB in a murine model of acute viral myocarditis. In a coxsackievirus B3 murine myocarditis model (Balb/c), effects of carvedilol on plasma noradrenaline, heart rate and blood pressure, myocardial histopathological changes and fibrosis, cardiomyocyte apoptosis, cardiac CREB and phosphorylated CREB, cytokine levels, and viral RNA were studied. RESULTS: The expression and phosphorylation of CREB were decreased with concomitant increase of IL-6 and TNF-α in murine coxsackievirus-induced acute viral myocarditis. The levels of IL-6 and TNF-α were correlated with the expression of CREB or phosphorylated CREB. Carvedilol increased the cardiac CREB expression and phosphorylation and decreased the plasma catecholamine levels and the production of IL-6 and TNF-α with amelioration of acute viral myocarditis. CONCLUSION: These results show that CREB may be involved in the pathophysiology of viral myocarditis and carvedilol exerts some of its beneficial effects by increasing the CREB expression and phosphorylation.

Molecular mechanisms associated with the antidepressant effects of the class I histone deacetylase inhibitor MS-275 in the rat ventrolateral orbital cortex.

Histone modifications mediated by histone acetylation are thought to play an important role in the pathogenesis and treatment of depression. Recent studies have revealed that histone deacetylase inhibitors (HDACis), such as sodium valproate (VPA) and MS-275, may be involved in the pathogenesis of depression and in the underpinnings of antidepressant therapeutic action in several brain regions, including the ventrolateral orbital cortex (VLO). In the present study, we investigated whether the class I histone deacetylase inhibitor MS-275 exerts antidepressant-like effects when infused bilaterally into the VLO of a rat, using the forced swimming test (FST) and tail suspension test (TST) as behavioral measures. We found that chronic intra-VLO infusion of MS-275 significantly reduced immobility time in the FST and TST compared with vehicle-treated controls, similar to the effects of systemically administered fluoxetine. These antidepressant-like effects of MS-275 are associated with an increase in H3 acetylation and elevated CREB and BDNF levels in the VLO. Our findings suggest the possibility that alterations in gene expression due to chromatin remodeling, including upregulation of CREB and BDNF, may be involved in the antidepressant-like effect of HDACis in the VLO.

LPS induces HMGB1 relocation and release by activating the NF-κB-CBP signal transduction pathway in the murine macrophage-like cell line RAW264.7.

BACKGROUND: High mobility group protein B1 (HMGB1) is an important late inflammatory mediator in sepsis. Understanding the mechanisms that regulate HMGB1 release from cells and their downstream signal transduction pathways may lead to the ability to develop anti-HMGB1 therapies to treat inflammation. MATERIALS AND METHODS: We stimulated murine macrophage-like RAW 264.7 cells with lipopolysaccharide (LPS) and LPS+ ethylpyruvate (EP) and examined the resulting HMGB1 expression and release. We also studied the expression of related signal transduction factors (NF-κB, p38 MAPK, and CBP). RESULTS AND CONCLUSION: Gene expression of HMGB1 mRNA in RAW264.7 cell showed no significant change at 0-18 h after stimulation with LPS, but increased significantly at 24, 36, and 48 h. HMGB1 mRNA expression in the LPS+EP group was significantly lower than in LPS alone. HMGB1 was distributed mainly in the nucleus; the cytoplasmic level was low before LPS stimulation. After stimulation with LPS, cytoplasmic HMGB1 increased gradually and plateaued at a high level at 12-48 h. Nuclear HMGB1 decreased gradually at 12-24 h, then increased, maintaining a comparatively high level at 36-48 h. EP prevented this pattern significantly. LPS induced p38 MAPK activation and NF-κB signal pathways first, followed by CBP activation. Activated CBP acetylated HMGB1 was stored in a crino-lysosome and secreted activated NF-κB resulted in increased transcription and synthesis of HMGB1, but the expression of up-regulated HMGB1 mRNA was delayed. Extracellular HMGB1 originated from early synthetic reserves present in the nucleus. New HMGB1 protein was synthesized in the nucleus and transferred into the cytoplasm, causing an increase in HMGB1 in the nucleus and cytoplasm. EP inhibits HMGB1 mRNA up-regulation and release from LPS- stimulated macrophages. The molecular function of EP is to attenuate the activation p38 MAPK, NF-κB, and CBP signaling pathways.

Self-inducible secretion of glucagon-like peptide-1 (GLP-1) that allows MIN6 cells to maintain insulin secretion and insure cell survival.

Based on the hypothesis that MIN6 cells could produce glucagon-like peptide-1 (GLP-1) to maintain cell survival, we analyzed the effects of GLP-1 receptor agonist, exendin-4 (Ex4), and antagonist, exendin-(9-39) (Ex9) on cell function and cell differentiation. MIN6 cells expressed proglucagon mRNAs and produced GLP-1, which was accelerated by Ex4 and suppressed by Ex9. Moreover, Ex4 further enhanced glucose-stimulated GLP-1 secretion, suggesting autocrine loop-contributed amplification of the GLP-1 signal. Ex4 up-regulated cell differentiation- and cell function-related CREBBP, Pdx-1, Pax6, proglucagon, and PC1/3 gene expressions. The confocal laser scanning images revealed that GLP-1 positive cells were dominant in the early stage of cells, but positive for insulin were more prominent in the mature stage of cells. Ex4 accelerated cell viability, while Ex9 and anti-GLP-1 receptor antibody enhanced cell apoptosis. MIN6 cells possess a mechanism of GLP-1 signal amplification in an autocrine fashion, by which the cells maintained insulin production and cell survival.

Gene expression profiles in BCL11B-siRNA treated malignant T cells.

BACKGROUND: Downregulation of the B-cell chronic lymphocytic leukemia (CLL)/lymphoma11B (BCL11B) gene by small interfering RNA (siRNA) leads to growth inhibition and apoptosis of the human T-cell acute lymphoblastic leukemia (T-ALL) cell line Molt-4. To further characterize the molecular mechanism, a global gene expression profile of BCL11B-siRNA -treated Molt-4 cells was established. The expression profiles of several genes were further validated in the BCL11B-siRNA -treated Molt-4 cells and primary T-ALL cells. RESULTS: 142 genes were found to be upregulated and 109 genes downregulated in the BCL11B-siRNA -treated Molt-4 cells by microarray analysis. Among apoptosis-related genes, three pro-apoptotic genes, TNFSF10, BIK, BNIP3, were upregulated and one anti-apoptotic gene, BCL2L1 was downregulated. Moreover, the expression of SPP1 and CREBBP genes involved in the transforming growth factor (TGF-ß) pathway was down 16-fold. Expression levels of TNFSF10, BCL2L1, SPP1, and CREBBP were also examined by real-time PCR. A similar expression pattern of TNFSF10, BCL2L1, and SPP1 was identified. However, CREBBP was not downregulated in the BLC11B-siRNA -treated Molt-4 cells. CONCLUSION: BCL11B-siRNA treatment altered expression profiles of TNFSF10, BCL2L1, and SPP1 in both Molt-4 T cell line and primary T-ALL cells.

Functional phosphosite screening for targeted protein-protein interactions by combining phosphoproteomics strategies and mammalian two-hybrid assays.

An effective new platform for phosphosite mapping and subsequent functional screening was developed to analyze the targeted protein-protein interactions of p300 and CBP with ß-catenin. Two novel functional phosphosites, Ser12 of p300 and Ser92 of CBP, were revealed to modulate p300/ß-catenin and CBP/ß-catenin interactions, respectively.

Presenilin controls CBP levels in the adult Drosophila central nervous system.

BACKGROUND: Dominant mutations in both human Presenilin (Psn) genes have been correlated with the formation of amyloid plaques and development of familial early-onset Alzheimer's disease (AD). However, a definitive mechanism whereby plaque formation causes the pathology of familial and sporadic forms of AD has remained elusive. Recent discoveries of several substrates for Psn protease activity have sparked alternative hypotheses for the pathophysiology underlying AD. CBP (CREB-binding protein) is a haplo-insufficient transcriptional co-activator with histone acetly-transferase (HAT) activity that has been proposed to be a downstream target of Psn signaling. Individuals with altered CBP have cognitive deficits that have been linked to several neurological disorders. METHODOLOGY/PRINCIPAL FINDINGS: Using a transgenic RNA-interference strategy to selectively silence CBP, Psn, and Notch in adult Drosophila, we provide evidence for the first time that Psn is required for normal CBP levels and for maintaining specific global acetylations at lysine 8 of histone 4 (H4K8ac) in the central nervous system (CNS). In addition, flies conditionally compromised for the adult-expression of CBP display an altered geotaxis behavior that may reflect a neurological defect. CONCLUSIONS/SIGNIFICANCE: Our data support a model in which Psn regulates CBP levels in the adult fly brain in a manner that is independent of Notch signaling. Although we do not understand the molecular mechanism underlying the association between Psn and CBP, our results underscore the need to learn more about the basic relationship between Psn-regulated substrates and essential functions of the nervous system.

Analysis of the expression pattern of the BCL11B gene and its relatives in patients with T-cell acute lymphoblastic leukemia.

BACKGROUND: In a human T-cell acute lymphoblastic leukemia (T-ALL) cell line (Molt-4), siRNA-mediated suppression of BCL11B expression was shown to inhibit proliferation and induce apoptosis, functions which may be related to genes involved in apoptosis (such as TNFSF10 and BCL2L1) and TGF-ß pathways (such as SPP1and CREBBP). METHODS: The expression levels of the above mentioned genes and their correlation with the BCL11B gene were analyzed in patients with T-ALL using the TaqMan and SYBR Green I real-time polymerase chain reaction technique. RESULTS: Expression levels of BCL11B, BCL2L1, and CREBBP mRNA in T-ALL patients were significantly higher than those from healthy controls (P<0.05). In T-ALL patients, the BCL11B expression level was negatively correlated with the BCL2L1 expression level (rs=-0.700; P<0.05), and positively correlated with the SPP1 expression level (rs=0.683; P<0.05). In healthy controls, the BCL11B expression level did not correlate with the TNFSF10, BCL2L1, SPP1, or CREBBP expression levels. CONCLUSIONS: Over-expression of BCL11B might play a role in anti-apoptosis in T-ALL cells through up-regulation of its downstream genes BCL2L1 and CREBBP.

Spatial memory consolidation is associated with induction of several lysine-acetyltransferase (histone acetyltransferase) expression levels and H2B/H4 acetylation-dependent transcriptional events in the rat hippocampus.

Numerous genetic studies have shown that the CREB-binding protein (CBP) is an essential component of long-term memory formation, through its histone acetyltransferase (HAT) function. E1A-binding protein p300 and p300/CBP-associated factor (PCAF) have also recently been involved in memory formation. By contrast, only a few studies have reported on acetylation modifications during memory formation, and it remains unclear as to how the system is regulated during this dynamic phase. We investigated acetylation-dependent events and the expression profiles of these HATs during a hippocampus-dependent task taxing spatial reference memory in the Morris water maze. We found a specific increase in H2B and H4 acetylation in the rat dorsal hippocampus, while spatial memory was being consolidated. This increase correlated with the degree of specific acetylated histones enrichment on some memory/plasticity-related gene promoters. Overall, a global increase in HAT activity was measured during this memory consolidation phase, together with a global increase of CBP, p300, and PCAF expression. Interestingly, these regulations were altered in a model of hippocampal denervation disrupting spatial memory consolidation, making it impossible for the hippocampus to recruit the CBP pathway (CBP regulation and acetylated-H2B-dependent transcription). CBP has long been thought to be present in limited concentrations in the cells. These results show, for the first time, that CBP, p300, and PCAF are dynamically modulated during the establishment of a spatial memory and are likely to contribute to the induction of a specific epigenetic tagging of the genome for hippocampus-dependent (spatial) memory consolidation. These findings suggest the use of HAT-activating molecules in new therapeutic strategies of pathological aging, Alzheimer's disease, and other neurodegenerative disorders.

Kaposi's sarcoma-associated herpesvirus viral protein kinase interacts with RNA helicase a and regulates host gene expression.

RNA helicase A (RHA) containing the DExH motif is a human homolog of maleless protein that regulates expression of genes located in the Drosophila X chromosome during dosage compensation. RHA exerts helicase activity that unwinds double-stranded RNA and DNA to a single-strand form. The protein acts as a bridging factor mediating interactions of CBP/p300 and RNA pol II, and consequently affects gene expression. Kaposi's sarcoma-associated herpesvirus (KSHV) is a member of the gamma-herpesvirus subfamily that causes several disorders. The majority of herpesviruses commonly encode predicted viral protein kinases. KSHV open reading frame 36 (ORF36) codes for protein kinase domains, and functions as a serine/threonine protein kinase. KSHV ORF36 is classified as a late gene, as it is expressed during lytic replication and localized in the nuclei of KSHV-infected cells. Recent studies show that viral protein kinase (vPK) interacts with cellular proteins. In this study, we determined the cellular localization of vPK in KSHV-infected BCBL-1 cells using confocal microscopy. Proteomic analysis indicates that cellular proteins interacted with vPK, and co-immunoprecipitation reactions further reveal interactions between vPK and RHA. Moreover, KSHV vPK appeared to regulate the transcriptional activation of Cre promoter, and plays an important role in cellular transcription of RHA.

Membrane-initiated estradiol signaling increases tyrosine hydroxylase promoter activity with ER alpha in PC12 cells.

Tyrosine hydroxylase (TH) promoter activity is induced by 17beta-estradiol (E(2)) in PC12 cells expressing estradiol receptor-alpha (ERalpha) requiring a cAMP/calcium response element (CRE/CaRE) at -45. To examine whether membrane-initiated estradiol signaling is underlying this induction, cells co-transfected with TH reporter construct and ERalpha expression vector were exposed to membrane-impermeant estradiol conjugate (beta-estradiol-6-(O-carboxy-methyl) oxime-bovine serum albumin, E(2)BSA). TH promoter activity was elevated by E(2)BSA in dose- and time-dependent manner. E(2)BSA also elicited rapid phosphorylation of CRE binding protein (CREB) and increased CRE-driven promoter activity. Over-expression of dominant negative forms of CREB, with mutations in DNA binding or phosphorylation site, prevented TH promoter response to E(2)BSA. Pre-treatment with protein kinase A (PKA) and MEK inhibitors reduced E(2) dependent phosphorylation of CREB and ERK, and also decreased induction of TH promoter activity by E(2) or E(2)BSA. Blocking S-palmitoylation of ERalpha with C451A mutation and/or pre-treatment with 2-Bromopalmitate did not prevent but instead enhanced E(2) or E(2)BSA-elicited induction of TH promoter activity. These findings reveal, for the first time, that estradiol induction of TH gene transcription with ERalpha in PC12 cells involves membrane-initiated estradiol signaling, rapid activation of dual PKA/MEK signaling pathways, leading to CREB phosphorylation, acting at CRE/CaRE. The data demonstrate possible mechanism whereby estradiol affects catecholaminergic systems in vivo.