Ser2 Phosphorylation Research Articles

Nitric oxide/cyclic guanosine monophosphate inducers sodium nitroprusside and L-arginine inhibit the proliferation of gastric cancer cells via the activation of type II cyclic guanosine monophosphate-dependent protein kinase.

Nitric oxide (NO) may activate soluble guanylyl cyclase (sGC), resulting in the increase of intracellular cyclic guanosine monophosphate (cGMP), a key molecule in the activation of type II cGMP-dependent protein kinase (PKG II). In our previous study, the membrane-permeable cGMP analogue 8-pCPT-cGMP was used to activate PKG II. The aim of the present study was to investigate whether NO/sGC-induced endogenous cGMP is able to activate PKG II and induce the corresponding effects. In the AGS gastric cancer cell line, the expression of PKG II was increased by infecting the cells with an adenoviral construct encoding PKG II cDNA (Ad-PKG II) and the activation of PKG II was induced by 8-pCPT-cGMP (positive control), the NO donor sodium nitroprusside (SNP) and the NO precursor L-arginine. ELISA was performed to detect the concentration of cGMP in AGS cells and the Cell Counting Kit-8 was used to analyze the proliferation of differently treated cells. Western blot analysis was used to detect the expression and phosphorylation of associated proteins. The results demonstrated that the level of cGMP was increased in cells treated with the NO donor or precursor. There was an obvious increase of Ser239 phosphorylation of the vasodilator-stimulated phosphoprotein, representing the increase in the activity of PKG II. The epidermal growth factor (EGF)-induced proliferation of AGS cells was inhibited by infection with Ad-PKG II and treatment with SNP or L-arginine. In addition, EGF-induced tyrosine phosphorylation of the EGF receptor (EGFR) and tyrosine/serine phosphorylation of extracellular signal-regulated kinase (ERK) were also inhibited by infection with Ad-PKG II and treatment with the NO donor or precursor. These data indicated that NO donors and precursors may activate the expression of PKG II, thereby blocking EGF-initiated signaling of the mitogen-activated protein kinase/ERK pathway and inhibiting EGF-induced proliferative activity through preventing the phosphorylation of EGFR at tyr1068.

Fisetin Ameliorated Photodamage by Suppressing the Mitogen-Activated Protein Kinase/Matrix Metalloproteinase Pathway and Nuclear Factor-κB Pathways.

Ultraviolet (UV) irradiation is one of the most important extrinsic factors contributing to skin photodamage. After UV irradiation, a series of signal transductions in the skin will be activated, leading to inflammatory response and photoaged skin. In this study, fisetin, a flavonol that exists in fruits and vegetables, was investigated for its photoprotective effects. The results revealed that 5-25 μM fisetin inhibits cyclooxygenase-2 (COX-2) and matrix metalloproteinase (MMP)-1, MMP-3, MMP-9 expression induced by ultraviolet B (UVB) irradiation in human skin fibroblasts. In addition, fisetin suppressed UVB-induced collagen degradation. With regard to its effect on upper-stream signal transduction, we found that fisetin reduced the expression of ultraviolet (UV)-induced ERK, JNK, and p38 phosphorylation in the mitogen-activated protein kinase (MAP kinase) pathway. Furthermore, fisetin reduced inhibitor κB (IκB) degradation and increased the amount of p65, which is a major subunit of nuclear factor-κB (NF-κB), in cytoplasm. It also suppressed NF-κB translocated to the nucleus and inhibited cAMP response element-binding protein (CREB) Ser-133 phosphorylation level in the phosphoinositide 3-kinase/protein kinase B/CREB (PI3K/AKT/CREB) pathway. Finally, fisetin inhibited UV-induced intracellular reactive oxygen species (ROS), prostaglandin E2 (PGE2), and nitric oxide (NO) generation. The mentioned effects and mechanisms suggest that fisetin can be used in the development of photoprotective agents.

SCP1 regulates c-Myc stability and functions through dephosphorylating c-Myc Ser62.

Serine 62 (Ser62) phosphorylation affects the c-Myc protein stability in cancer cells. However, the mechanism for dephosphorylating c-Myc is not well understood. In this study, we identified carboxyl-terminal domain RNA polymerase II polypeptide A small phosphatase 1 (SCP1) as a novel phosphatase specifically dephosphorylating c-Myc Ser62. Ectopically expressed SCP1 strongly dephosphorylated c-Myc Ser62, destabilized c-Myc protein and suppressed c-Myc transcriptional activity. Knockdown of SCP1 increased the c-Myc protein levels in liver cancer cells. SCP1 interacted with c-Myc both in vivo and in vitro. In addition, Ser245 at the C-terminus of SCP1 was essential for its phosphatase activity towards c-Myc. Functionally, SCP1 negatively regulated the cancer cell proliferation. Collectively, our findings indicate that SCP1 is a potential tumor suppressor for liver cancers through dephosphorylating c-Myc Ser62.

Pcf11 orchestrates transcription termination pathways in yeast.

In Saccharomyces cerevisiae, short noncoding RNA (ncRNA) generated by RNA polymerase II (Pol II) are terminated by the NRD complex consisting of Nrd1, Nab3, and Sen1. We now show that Pcf11, a component of the cleavage and polyadenylation complex (CPAC), is also generally required for NRD-dependent transcription termination through the action of its C-terminal domain (CTD)-interacting domain (CID). Pcf11 localizes downstream from Nrd1 on NRD terminators, and its recruitment depends on Nrd1. Furthermore, mutation of the Pcf11 CID results in Nrd1 retention on chromatin, delayed degradation of ncRNA, and restricted Pol II CTD Ser2 phosphorylation and Sen1-Pol II interaction. Finally, the pcf11-13 and sen1-1 mutant phenotypes are very similar, as both accumulate RNA:DNA hybrids and display Pol II pausing downstream from NRD terminators. We predict a mechanism by which the exchange of Nrd1 and Pcf11 on chromatin facilitates Pol II pausing and CTD Ser2-P phosphorylation. This in turn promotes Sen1 activity that is required for NRD-dependent transcription termination in vivo.

Effect of all-transretinoic acid on differentiation of cancer stem cells in colorectal cancer

Objective To detect the promoting effects of all- trans retinoic acid(ATRA)on differentiation of cancer stem cells(CSCs)of colorectal cancer and the mechanism. Methods In order to enrich CSCs, magnetic activated cell sorting(MACS) was used to purify CD133+ CSCs from SW620 cell line. The proportion of CD133+ cells in purified CSCs was measured after treatment with 1.0×10-6 mol/ L ATRA or dimethylsurfoxide(DMSO)for 3 days by flow cytometry. The inhibitory effect of selfrenewing ability of CSCs induced by 1.0×10-6 mol/L ATRA was evaluated by sphere formation assay. Phosphorylation and degradation of β- catenin induced by 1.0×10-6 mol/L ATRA was confirmed by Western blotting. Results CD133+ SW620 cells effectively enrich CSCs. The rate of tumor spheres of CD133+ and CD133- cells was(51.00±7.67)% and(4.58±2.52)%, respectively (P< 0.01). After treatment with 1.0×10-6 mol/L ATRA, the proportion of CD133+ cells was (47.90±7.87)%,and that in the control group was(95.33±2.24)%, respectively(P< 0.01). Interestingly, the rate of tumor spheres was decreased from(53.66±6.03)% to(19.33±2.52)% (P< 0.01). Western blotting indicated that ATRA induced degradation of β- catenin by Thr41/ Ser45 phosphorylation. Conclusion ATRA induces differentiation of colorectal CSCs, and inhibits their self- renewing ability by suppression of Wnt/β- catenin pathway. Key words: All-trans retinoic acid; Colorectal cancer; Cancer stem cells; Cell differentiation

Interactome Dynamics of the Transcription Elongation Complex Revealed by Quantitative Proteomics

At each stage of transcription, RNA polymerase is regulated by a host of dynamically recruited factors. Many stages are associated with the phosphorylation state of the C‐terminal domain of the largest Pol II subunit, which consists of the peptide repeat Y1S2P3T4S5P6S7. Factors that co‐purify with a particular phosphoisoform of the CTD likely act at the associated stage of transcription and are recruited either through direct phospho‐CTD binding, other Pol II subunits and regulatory factors, the nascent RNA or the DNA template. To probe the dynamic composition of the transcription elongation complex, the interactome of each CTD phosphoisoform in S. cerevisiae was determined using a sequential immunoprecipitaiton strategy followed by label‐free mass spectrometry. Our data identify an average of 80 statistically significant Pol II interactors per phoshoisoform, many of which display a preference for purifying with specific isoforms. For example, Spt4, a transcription elongation factor, is highly enriched in the Ser5P dataset, suggesting that it acts in the 5′ region of genes, where Ser5 phosphorylation is most abundant. Indeed, our analysis of global Pol II density in spt4 deletion mutants by native elongating transcript sequencing (NET‐seq) revealed strong transcriptional defects in the 5′ region. Other factors, such as the termination factor Rai1, interacted with all isoforms. NET‐seq analysis of a yeast strain lacking rai1 revealed major transcriptional defects across gene bodies and in termination. This defect was seen in both sense and anti‐sense transcripts, indicating that Rai1 has a previously unappreciated general role in regulating transcription. Taken together these data provide a network of Pol II interacting factors that function to regulate transcription.

The union of transcription and mRNA processing: 20 years of coupling.

The union of transcription and mRNA processing: 20 years of coupling.

PCTAIRE1/PCTK1/CDK16: a new oncotarget?

PCTAIRE1/PCTK1/CDK16: a new oncotarget?

P-TEFb, the super elongation complex and mediator regulate a subset of non-paused genes during early Drosophila embryo development.

Positive Transcription Elongation Factor b (P-TEFb) is a kinase consisting of Cdk9 and Cyclin T that releases RNA Polymerase II (Pol II) into active elongation. It can assemble into a larger Super Elongation Complex (SEC) consisting of additional elongation factors. Here, we use a miRNA-based approach to knock down the maternal contribution of P-TEFb and SEC components in early Drosophila embryos. P-TEFb or SEC depletion results in loss of cells from the embryo posterior and in cellularization defects. Interestingly, the expression of many patterning genes containing promoter-proximal paused Pol II is relatively normal in P-TEFb embryos. Instead, P-TEFb and SEC are required for expression of some non-paused, rapidly transcribed genes in pre-cellular embryos, including the cellularization gene Serendipity-α. We also demonstrate that another P-TEFb regulated gene, terminus, has an essential function in embryo development. Similar morphological and gene expression phenotypes were observed upon knock down of Mediator subunits, providing in vivo evidence that P-TEFb, the SEC and Mediator collaborate in transcription control. Surprisingly, P-TEFb depletion does not affect the ratio of Pol II at the promoter versus the 3’ end, despite affecting global Pol II Ser2 phosphorylation levels. Instead, Pol II occupancy is reduced at P-TEFb down-regulated genes. We conclude that a subset of non-paused, pre-cellular genes are among the most susceptible to reduced P-TEFb, SEC and Mediator levels in Drosophila embryos.

S-nitrosylation of cofilin-1 mediates estradiol-17β-stimulated endothelial cytoskeleton remodeling.

Rapid nitric oxide (NO) production via endothelial NO synthase (eNOS) activation represents a major signaling pathway for the cardiovascular protective effects of estrogens; however, the pathways after NO biosynthesis that estrogens use to function remain largely unknown. Covalent adduction of a NO moiety to cysteines, termed S-nitrosylation (SNO), has emerged as a key route for NO to directly regulate protein function. Cofilin-1 (CFL1) is a small actin-binding protein essential for actin dynamics and cytoskeleton remodeling. Despite being identified as a major SNO protein in endothelial cells, whether SNO regulates CFL-1 function is unknown. We hypothesized that estradiol-17β (E2β) stimulates SNO of CFL1 via eNOS-derived NO and that E2β-induced SNO-CFL1 mediates cytoskeleton remodeling in endothelial cells. Point mutation studies determined Cys80 as the primary SNO site among the 4 cysteines (Cys39/80/139/147) in CFL1. Substitutions of Cys80 with Ala or Ser were used to prepare the SNO-mimetic/deficient (C80A/S) CFL1 mutants. Recombinant wild-type (wt) and mutant CFL1 proteins were prepared; their actin-severing activity was determined by real-time fluorescence imaging analysis. The activity of C80A CFL1 was enhanced to that of the constitutively active S3/A CFL1, whereas the other mutants had no effects. C80A/S mutations lowered Ser3 phosphorylation. Treatment with E2β increased filamentous (F)-actin and filopodium formation in endothelial cells, which were significantly reduced in cells overexpressing wt-CFL. Overexpression of C80A, but not C80S, CFL1 decreased basal F-actin and further suppressed E2β-induced F-actin and filopodium formation compared with wt-CFL1 overexpression. Thus, SNO(Cys80) of cofilin-1 via eNOS-derived NO provides a novel pathway for mediating estrogen-induced endothelial cell cytoskeleton remodeling.

Serine 133 phosphorylation is not required for hippocampal CREB-mediated transcription and behavior.

The cAMP response element (CRE)-binding protein, CREB, is a transcription factor whose activity in the brain is critical for long-term memory formation. Phosphorylation of Ser133 in the kinase-inducible domain (KID), that in turn leads to the recruitment of the transcriptional coactivator CREB-binding protein (CBP), is thought to mediate the activation of CREB. However, the importance of phosphorylation for CREB binding to DNA and subsequent gene transcription in vivo is controversial. To definitively address the role of CREB phosphorylation in gene transcription and learning and memory, we derived mutant mice lacking the Ser133 phosphorylation site. These mice exhibit normal CREB-mediated gene transcription for a number of genes implicated in learning and memory processes. Furthermore these mice have no deficits in hippocampus- or striatum-dependent learning. Strikingly, our findings show that CREB phosphorylation at Ser133 is not necessary for CREB binding to CRE sites, CREB-mediated transcription, or CREB-mediated behavioral phenotypes associated with learning and memory.

Methotrexate-mediated activation of an AMPK-CREB-dependent pathway: a novel mechanism for vascular protection in chronic systemic inflammation

AimsPremature cardiovascular events complicate chronic inflammatory conditions. Low-dose weekly methotrexate (MTX), the most widely used disease-modifying drug for rheumatoid arthritis (RA), reduces disease-associated cardiovascular mortality. MTX increases intracellular accumulation of...

HDAC6 inhibitors modulate Lys49 acetylation and membrane localization of β-catenin in human iPSC-derived neuronal cells.

We examined the effects of isoform-specific histone deacetylase (HDAC) inhibitors on β-catenin posttranslational modifications in neural progenitor cells (NPCs) derived from human induced pluripotent stem cells (iPSCs). β-catenin is a multifunctional protein with important roles in the developing and adult central nervous system. Activation of the Wnt pathway results in stabilization and nuclear translocation of β-catenin, resulting in activation of multiple target genes. In addition, β-catenin forms a complex with cadherins at the plasma membrane as part of the adherens junctions. The N-terminus of β-catenin has phosphorylation, ubiquitination, and acetylation sites that regulate its stability and signaling. In the absence of a Wnt signal, Ser33, Ser37, and Thr41 are constitutively phosphorylated by glycogen synthase kinase 3β (GSK3β). β-Catenin phosphorylated at these sites is recognized by β-transducin repeat-containing protein (βTrCP), which results in ubiquitination and degradation by the ubiquitin-proteasome pathway. The N-terminal regulatory domain of β-catenin also includes Ser45, a phosphorylation site for Casein Kinase 1α (CK1α) and Lys49, which is acetylated by the acetyltransferase p300/CBP-associated factor (PCAF). The relevance of Lys49 acetylation and Ser45 phosphorylation to the function of β-catenin is an active area of investigation. We find that HDAC6 inhibitors increase Lys49 acetylation and Ser45 phosphorylation but do not affect Ser33, Ser37, and Thr41 phosphorylation. Lys49 acetylation results in decreased ubiquitination of β-catenin in the presence of proteasome inhibition. While increased Lys49 acetylation does not affect total levels of β-catenin, it results in increased membrane localization of β-catenin.

EIF2α Confers Cellular Tolerance to S. aureus α-Toxin.

We report on the role of conserved stress–response pathways for cellular tolerance to a pore forming toxin. First, we observed that small molecular weight inhibitors including of eIF2α-phosphatase, jun-N-terminal kinase (JNK), and PI3-kinase sensitized normal mouse embryonal fibroblasts (MEFs) to the small pore forming S. aureus α-toxin. Sensitization depended on expression of mADAM10, the murine ortholog of a proposed high-affinity receptor for α-toxin in human cells. Similarly, eIF2αS51A/S51A MEFs, which harbor an Ala knock-in mutation at the regulated Ser51 phosphorylation site of eukaryotic translation initiation factor 2α, were hyper-sensitive to α-toxin. Inhibition of translation with cycloheximide did not mimic the tolerogenic effect of eIF2α-phosphorylation. Notably, eIF2α-dependent tolerance of MEFs was toxin-selective, as wild-type MEFs and eIF2αS51A/S51A MEFs exhibited virtually equal sensitivity to Vibrio cholerae cytolysin. Binding of S. aureus α-toxin to eIF2αS51A/S51A MEFs and toxicity in these cells were enhanced as compared to wild-type cells. This led to the unexpected finding that the mutant cells carried more ADAM10. Because basal phosphorylation of eIF2α in MEFs required amino acid deprivation-activated eIF2α-kinase 4/GCN2, the data reveal that basal activity of this kinase mediates tolerance of MEFs to α-toxin. Further, they suggest that modulation of ADAM10 is involved. During infection, bacterial growth may cause nutrient shortage in tissues, which might activate this response. Tolerance to α-toxin was robust in macrophages and did not depend on GCN2. However, JNKs appeared to play a role, suggesting differential cell type and toxin selectivity of tolerogenic stress responses. Understanding their function or failure will be important to comprehend anti-bacterial immune responses.

Serine phosphorylation of vasodilator-stimulated phosphoprotein (VASP) regulates colon cancer cell survival and apoptosis

AimsIn colon cancer, disease recurrence and death are associated with abnormal tumor cell survival. Vasodilator-stimulated phosphoprotein (VASP) is an actin binding protein regulating cell shape and polarity through the F-actin cytoskeleton, whose activity is controlled by cAMP-dependent phosphorylation at Ser157 and cGMP-dependent phosphorylation at Ser239. This study examined the role of differential VASP Ser phosphorylation in regulating cell survival and apoptosis in human colon carcinoma cells. Main methodsSelective inhibition of VASP Ser157 or Ser239 phosphorylation in colon cancer cells was performed with specific phosphomutant constructs. F-actin organization was examined by confocal microscopy, and the balance of cell survival and death assessed by measuring acridine orange and ethidium bromide staining, caspase-3 and BAD-pS112 expression and DNA fragmentation. Key findingsIn human colon carcinoma cells suppression of VASP Ser157 phosphorylation reduced F-actin content and survival and increased apoptosis, while inhibition of VASP Ser239 phosphorylation increased F-actin content and survival and reduced cell death. Also, while 8Br-cAMP induced VASP Ser157 phosphorylation and reduced cell death, treatments with 8CPT-cGMP elevated VASP Ser239 phosphorylation and promoted apoptosis. SignificanceThese findings suggest that differential VASP Ser phosphorylation represents a unique therapeutic target to control cell survival and death behavior in colon cancer. In particular, pharmacological manipulation of VASP Ser phosphorylation could be exploited to affect the malignant actin cytoskeleton and induce apoptosis in colorectal cancer cells.

Preconditioning Human Mesenchymal Stem Cells with a Low Concentration of BMP2 Stimulates Proliferation and Osteogenic Differentiation In Vitro.

Clinical trials using bone morphogenetic protein-2 (BMP2) for bone reconstruction have shown promising results. However, the relatively high concentration needed to be effective raises concerns for efficacy and safety. The aim of this study was to investigate the osteogenic effect of an alternative treatment strategy in which human bone marrow–derived mesenchymal stem cells (hMSCs) are preconditioned with low concentrations of BMP2 for a short time in vitro. hMSCs in suspension were stimulated for 15 min with 10 and 20 ng/mL of BMP2. After the BMP2 was removed, the cells were seeded and cultured in osteogenic medium. The effects of preconditioning were analyzed with regard to proliferation and expression of osteogenic markers at both gene and protein level. The results were compared to those from cultures with continuous BMP2 stimulation. A significant increase in proliferation was seen with both precondition and continuous stimulation with BMP2, with no difference between the treatments. Preconditioning with BMP2 significantly increased gene expression of RUNX2, COLI, ALP, and OC, and protein levels of COLI and ALP. This was not found with continuous stimulation. The role of preconditioning with BMP2 in osteogenesis was validated by findings of increased gene expression of SMAD1 and an increase in dual phosphorylation of ser 463 and ser 465 in the SMAD 1/5/8 pathway. We concluded that preconditioning hMSCs with BMP2 stimulates osteogenesis: proliferation with matrix secretion and matrix maturation of hMSCs. This implies that preconditioning with BMP2 might be more effective at inducing proliferation and osteogenic differentiation of hMSCs than continuous stimulation. Preconditioning with BMP2 could benefit the clinical application of BMP2 since side effects from high-dose treatments could be avoided.

Role of non-genomic androgen signalling in suppressing proliferation of fibroblasts and fibrosarcoma cells.

The functions of androgen receptor (AR) in stromal cells are still debated in spite of the demonstrated importance of these cells in organ development and diseases. Here, we show that physiological androgen concentration (10 nM R1881 or DHT) fails to induce DNA synthesis, while it consistently stimulates cell migration in mesenchymal and transformed mesenchymal cells. Ten nanomolar R1881 triggers p27 Ser10 phosphorylation and its stabilization in NIH3T3 fibroblasts. Activation of Rac and its downstream effector DYRK 1B is responsible for p27 Ser10 phosphorylation and cell quiescence. Ten nanomolar androgen also inhibits transformation induced by oncogenic Ras in NIH3T3 fibroblasts. Overexpression of an AR mutant unable to interact with filamin A, use of a small peptide displacing AR/filamin A interaction, and filamin A knockdown indicate that the androgen-triggered AR/filamin A complex regulates the pathway leading to p27 Ser10 phosphorylation and cell cycle arrest. As the AR/filamin A complex is also responsible for migration stimulated by 10 nM androgen, our report shows that the androgen-triggered AR/filamin A complex controls, through Rac 1, the decision of cells to halt cell cycle and migration. This study reveals a new and unexpected role of androgen/AR signalling in coordinating stromal cell functions.

Rpb4/7 facilitates RNA polymerase II CTD dephosphorylation.

The Rpb4 and Rpb7 subunits of eukaryotic RNA polymerase II (RNAPII) participate in a variety of processes from transcription, DNA repair, mRNA export and decay, to translation regulation and stress response. However, their mechanism(s) of action remains unclear. Here, we show that the Rpb4/7 heterodimer in Saccharomyces cerevisiae plays a key role in controlling phosphorylation of the carboxy terminal domain (CTD) of the Rpb1 subunit of RNAPII. Proper phosphorylation of the CTD is critical for the synthesis and processing of RNAPII transcripts. Deletion of RPB4, and mutations that disrupt the integrity of Rpb4/7 or its recruitment to the RNAPII complex, increased phosphorylation of Ser2, Ser5, Ser7 and Thr4 within the CTD. RPB4 interacted genetically with genes encoding CTD phosphatases (SSU72, FCP1), CTD kinases (KIN28, CTK1, SRB10) and a prolyl isomerase that targets the CTD (ESS1). We show that Rpb4 is important for Ssu72 and Fcp1 phosphatases association, recruitment and/or accessibility to the CTD, and that this correlates strongly with Ser5P and Ser2P levels, respectively. Our data also suggest that Fcp1 is the Thr4P phosphatase in yeast. Based on these and other results, we suggest a model in which Rpb4/7 helps recruit and potentially stimulate the activity of CTD-modifying enzymes, a role that is central to RNAPII function.

Epigenetic regulation of the NR4A orphan nuclear receptor NOR1 by histone acetylation

The nuclear receptor NOR1 is an immediate-early response gene implicated in the transcriptional control of proliferation. Since the expression level of NOR1 is rapidly induced through cAMP response element binding (CREB) protein-dependent promoter activation, we investigated the contribution of histone acetylation to this transient induction. We demonstrate that NOR1 transcription is induced by histone deacetylase (HDAC) inhibition and by depletion of HDAC1 and HDAC3. HDAC inhibition activated the NOR1 promoter, increased histone acetylation and augmented the recruitment of phosphorylated CREB to the promoter. Furthermore, HDAC inhibition increased Ser133 phosphorylation of CREB and augmented NOR1 protein stability. These data outline previously unrecognized mechanisms of NOR1 regulation and illustrate a key role for histone acetylation in the rapid induction of NOR1.

Combinatorial regulation of a signal-dependent activator by phosphorylation and acetylation.

In the fasted state, increases in catecholamine signaling promote adipocyte function via the protein kinase A-mediated phosphorylation of cyclic AMP response element binding protein (CREB). CREB activity is further up-regulated in obesity, despite reductions in catecholamine signaling, where it contributes to the development of insulin resistance. Here we show that obesity promotes the CREB binding protein (CBP)-mediated acetylation of CREB at Lys136 in adipose. Under lean conditions, CREB acetylation was low due to an association with the energy-sensing NAD(+)-dependent deacetylase SirT1; amounts of acetylated CREB were increased in obesity, when SirT1 undergoes proteolytic degradation. Whereas CREB phosphorylation stimulated an association with the KIX domain of CBP, Lys136 acetylation triggered an interaction with the CBP bromodomain (BRD) that augmented recruitment of this coactivator to the promoter. Indeed, coincident Ser133 phosphorylation and Lys136 acetylation of CREB stimulated the formation of a ternary complex with the KIX and BRD domains of CBP by NMR analysis. As disruption of the CREB:BRD complex with a CBP-specific BRD inhibitor blocked effects of CREB acetylation on target gene expression, our results demonstrate how changes in nutrient status modulate cellular gene expression in response to hormonal signals.