p300/CBP-associated Factor Research Articles

Pharmacological blockade of PCAF ameliorates osteoarthritis development via dual inhibition of TNF-α-driven inflammation and ER stress.

BackgroundEpigenetic mechanisms have been reported to play key roles in osteoarthritis (OA) development. P300/CBP-associated factor (PCAF) is a member of the histone acetyltransferases, which exhibits a strong relationship with endoplasmic reticulum (ER) stress and transcription factor nuclear factor kappa B (NF-κB) signals. Salidroside, a natural histone acetylation inhibitor, showed its anti-inflammatory and anti-apoptotic effects in lipopolysaccharide (LPS)-stimulated microglia cells in our previous study. However, whether Sal has a protective effect against OA remains unknown, and its relationships to PCAF, NF-κB, and the ER stress pathway should be explored further.MethodsWe identified the role of PCAF in the pathogenesis of OA and determined the chondroprotective effect of Sal on both tumor necrosis factor alpha (TNF-α)-treated human chondrocytes and a destabilized medial meniscus (DMM) mouse OA model.FindingsWe found increased PCAF expression in human OA cartilage and TNF-α-driven chondrocytes. Meanwhile, silencing of PCAF attenuated nuclear p65 and C/EBP homologous protein levels in chondrocytes upon TNF-α stimulation. Furthermore, Sal was found to specifically bind to the inhibitory site of the PCAF protein structure, which subsequently reversed the TNF-α-induced activation of NF-κB signal and ER stress-related apoptosis in chondrocytes. In addition, the protective effect of Sal and its inhibitory effects on PCAF as well as inflammatory- and ER stress-related markers were also observed in the mouse DMM model.InterpretationPharmacological blockade of PCAF by Sal ameliorates OA development via inhibition of inflammation and ER stress, which makes Sal a promising therapeutic agents for the treatment of OA.

The effect of dietary garcinol supplementation on oxidative stability, muscle postmortem glycolysis and meat quality in pigs

The objective of this study was to evaluate the effects of dietary garcinol (0, 200, 400 and 600 mg/kg) on the growth performance, meat quality, postmortem glycolysis and antioxidative capacity of finishing pigs. Dietary garcinol increased pigs' average daily gain, pH 24h, a* and myoglobin content of longissimus dorsi (LM) (P < 0.05), and decreased feed/gain ratio, the L*24h, glycolytic potential, drip loss, shear force, and backfat depth (P < 0.05). The glutathione peroxidase (GPx), catalase (CAT) and total antioxidative capacity (T-AOC) were significantly increased by garcinol (P < 0.05), while the activity of lactate dehydrogenase (LDH) and malonaldehyde (MDA) content were decreased (P < 0.05). Moreover, garcinol decreased the p300/CBP-associated factor (PCAF) activity, the acetylation level and activities of glycolysis enzymes phosphoglycerate kinase 1 (PGK1), glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and 6-phosphofructo-2-kinase/fructose-2, 6-bisphosphatase-3 (PFKFB3) (P < 0.05). The results of this study showed that garcinol decreased postmortem glycolysis, and this may be due to the mechanism of decreasing glycolytic enzyme acetylation induced by PCAF. The present study indicates that garcinol can facilitate the growth performance of pigs and improve pork quality by changing postmortem glycolysis and antioxidative capacity.

K-Ras-PI3K regulates H3K56ac through PCAF to elevate the occurrence and growth of liver cancer.

H3 modification is related to a wide range of tumors, including liver cancer. The Ras passageway is actuated in human diseases. Thus, we investigated the roles of Ras in liver cancer cells via acetylation of H3K56. Ras-carrying G12V and Y40C site mutation was transfected into liver cancer cell lines SNU-475 and SK-Hep-1. Acetylation of H3K56 and phosphatidylinositol 3-kinase (PI3K), P300/CBP-associated factor (PCAF) and Mouse double minute 2 homolog (MDM2) was tested via western blot. Cell activity, colonies, and migration were tested via Cell Counting Kit-8, soft-agar colony formation, and Transwell experiment, respectively. Sirtuin 6 (SIRT6) and PCAF were tested via quantitative reverse transcription polymerase chain reaction (qRT-PCR). Chromatin immunoprecipitation was employed to test the relationship between Ras and downstream elements. Flow cytometry was employed to test cell cycle series. We found that RasG12V/Y40C transfection reduced the acetylation of H3K56 and activated phosphorylation of protein kinase B. H3K56Q (H3K56ac overexpression) suppressed cell activity, colonies, and migration. H3K56ac changed Ras downstream factors expression. RasG12V/Y40C bound to Ras-PI3K downstream elements' promoters. SIRT6 silencing raised H3K56ac and suppressed cell activity, migration and S phage cell percentage. SIRT6 silence transformed expression of downstream elements. PCAF and H3K56ac demonstrated the close current while MDM2 was conversed. In summary, the Ras-PI3K passageway promoted cell growth and metastasis via decreasing H3K56ac, in which MDM2-mediated PCAF was involved.

Acetylation of ezrin regulates membrane-cytoskeleton interaction underlying CCL18-elicited cell migration.

Ezrin, a membrane–cytoskeleton linker protein, plays an essential role in cell polarity establishment, cell migration, and division. Recent studies show that ezrin phosphorylation regulates breast cancer metastasis by promoting cancer cell survivor and promotes intrahepatic metastasis via cell migration. However, it was less characterized whether there are additional post-translational modifications and/or post-translational crosstalks on ezrin underlying context-dependent breast cancer cell migration and invasion. Here we show that ezrin is acetylated by p300/CBP-associated factor (PCAF) in breast cancer cells in response to CCL18 stimulation. Ezrin physically interacts with PCAF and is a cognate substrate of PCAF. The acetylation site of ezrin was mapped by mass spectrometric analyses, and dynamic acetylation of ezrin is essential for CCL18-induced breast cancer cell migration and invasion. Mechanistically, the acetylation reduced the lipid-binding activity of ezrin to ensure a robust and dynamic cycling between the plasma membrane and cytosol in response to CCL18 stimulation. Biochemical analyses show that ezrin acetylation prevents the phosphorylation of Thr567. Using atomic force microscopic measurements, our study revealed that acetylation of ezrin induced its unfolding into a dominant structure, which prevents ezrin phosphorylation at Thr567. Thus, these results present a previously undefined mechanism by which CCL18-elicited crosstalks between the acetylation and phosphorylation on ezrin control breast cancer cell migration and invasion. This suggests that targeting PCAF signaling could be a potential therapeutic strategy for combating hyperactive ezrin-driven cancer progression.

Ambient and supplemental magnetic fields promote myogenesis via a TRPC1-mitochondrial axis: evidence of a magnetic mitohormetic mechanism.

We show that both supplemental and ambient magnetic fields modulate myogenesis. A lone 10 min exposure of myoblasts to 1.5 mT amplitude supplemental pulsed magnetic fields (PEMFs) accentuated in vitro myogenesis by stimulating transient receptor potential (TRP)-C1-mediated calcium entry and downstream nuclear factor of activated T cells (NFAT)-transcriptional and P300/CBP-associated factor (PCAF)-epigenetic cascades, whereas depriving myoblasts of ambient magnetic fields slowed myogenesis, reduced TRPC1 expression, and silenced NFAT-transcriptional and PCAF-epigenetic cascades. The expression levels of peroxisome proliferator-activated receptor γ coactivator 1α, the master regulator of mitochondriogenesis, was also enhanced by brief PEMF exposure. Accordingly, mitochondriogenesis and respiratory capacity were both enhanced with PEMF exposure, paralleling TRPC1 expression and pharmacological sensitivity. Clustered regularly interspaced short palindromic repeats-Cas9 knockdown of TRPC1 precluded proliferative and mitochondrial responses to supplemental PEMFs, whereas small interfering RNA gene silencing of TRPM7 did not, coinciding with data that magnetoreception did not coincide with the expression or function of other TRP channels. The aminoglycoside antibiotics antagonized and down-regulated TRPC1 expression and, when applied concomitantly with PEMF exposure, attenuated PEMF-stimulated calcium entry, mitochondrial respiration, proliferation, differentiation, and epigenetic directive in myoblasts, elucidating why the developmental potential of magnetic fields may have previously escaped detection. Mitochondrial-based survival adaptations were also activated upon PEMF stimulation. Magnetism thus deploys an authentic myogenic directive that relies on an interplay between mitochondria and TRPC1 to reach fruition.-Yap, J. L. Y., Tai, Y. K., Fröhlich, J., Fong, C. H. H., Yin, J. N., Foo, Z. L., Ramanan, S., Beyer, C., Toh, S. J., Casarosa, M., Bharathy, N., Kala, M. P., Egli, M., Taneja, R., Lee, C. N., Franco-Obregón, A. Ambient and supplemental magnetic fields promote myogenesis via a TRPC1-mitochondrial axis: evidence of a magnetic mitohormetic mechanism.

PCAF regulates H3 phosphorylation and promotes autophagy in osteosarcoma cells

BackgroundOsteosarcoma is one of malignant cancer. Histone phosphorylation is common in tumors. We explored the effects of p300-CBP-associated factor (PCAF) and phosphorylation of H3S28 in osteosarcoma cancer cell autophagy. MethodsOsteosarcoma cancer cell lines were collected and/or transfected with full length PCAF or interference miRNAs to mimic or silence of PCAF expression. Immunoprecipitation assay and GST pull down was used to target targeting PCAF or H3S28ph. H3-/- SNU-C1 cells were transfected with H3WT- or H3S28F-expressing or enhanced green fluorescent protein (EGFP)-tagged LC3 plasmids, in which H3 was tagged with HA. An in vitro kinase activity assay was performed to test whether recombinant full-length PCAF could phosphorylate H3 in the site of S28. The functions on autophagy was detected by number of autophagosomes, number of EGFP-LC3, LC3-II/I, percentage of degradation and expression of autophagy associated gene (ATG). ResultsPCAF positively regulated H3S28ph in osteosarcoma cancer cells; Immunoprecipitation assay and GST pull down demonstrated that PCAF could interact directly with H3 in osteosarcoma cancer cells. In addition, silence of PCAF inhibited the number of autophagosomes, number of EGFP-LC3, LC3-II/I, percentage of degradation and expression of ATG. Moreover, H3S28A (H3S28 mutation) impaired the promoting autophagy effects of PCAF. The PCAF-H3S28ph axis promoted osteosarcoma cancer autophagy viatranscriptional regulation of ATG genes. ConclusionPCAF regulated H3S28 phosphorylation and their axis promotes autophagy in osteosarcoma cancer cells viatargeting ATG5 and ATG7.

Induction of NKG2D ligand expression on tumor cells by CD8+ T-cell engagement-mediated activation of nuclear factor-kappa B and p300/CBP-associated factor.

The ligands for the natural killer group 2 (NKG2D) protein render tumor cells susceptible to NKG2D-dependent immune cell attack. However, cancer cells escape from immune surveillance by downregulating NKG2D ligands. We previously discovered that engagement of activated CD8+ T cells and tumor cells induces NKG2D ligands on tumor cells, but the underlying mechanism remains to be defined. Both in vivo mouse tumor models and in vitro cell assays were performed to study the downstream signaling. Our results supported the notion that, upon engagement with the cognate receptors, CD137 ligand and CD40 initiates activation of nuclear factor-kappa B (NF-κB) signaling in tumor cells even in the absence of CD8+ T cells. Like tumor and CD8+ T cell contact-dependent NKG2D ligand induction, this CD137L/CD40-mediated signaling activation was associated with elevated levels of acetyltransferase P300/CBP-associated factor (PCAF), whereas inhibition of phosphorylated NF-κB abrogated PCAF induction. Although stimulation of CD137L/CD40-mediated signaling is vital, inflammatory cytokines, including interferon gamma (IFNγ) and TNFα, also facilitate NKG2D ligand–induced immune surveillance via both facilitating T cell chemotaxis and CD137L/CD40 induced NF-κB/PCAF activation. Collectively, our results unveil a novel mechanism of NKG2D ligand upregulation involving reverse signaling of CD40 and CD137L on tumor cells which, along with inflammatory cytokines IFNγ and TNFα, stimulate downstream NF-κB and PCAF activation. Understanding this mechanism may help in development of induced NKG2D ligand–dependent T cell therapy against cancers.

Pharmacological Blockade of PCAF Ameliorates Osteoarthritis Development via Dual Inhibition of TNF-α-Driven Inflammation and ER Stress

Epigenetic mechanisms have been reported to play key roles in osteoarthritis (OA) development. Here, we identified the role of histone acetyltransferase p300/CBP-associated factor (PCAF) in the pathogenesis of OA and determined that the chondroprotective effect of specific pharmacological inhibition of PCAF by a natural compound salidroside (Sal) in Tumor Necrosis Factor Alpha (TNF-α) treated human chondrocytes and a destabilized medial meniscus (DMM) mouse OA model.In vitro, we found increased PCAF expression and activity in human OA cartilage and TNF-α-driven chondrocytes. Meanwhile, silencing PCAF attenuates the nuclear p65 and C/EBP Homologous Protein (CHOP) level in human chondrocytes under TNF-α stimulation. Furthermore, Sal was found to bind the inhibitory site of PCAF protein structure specifically according to docking analysis, which subsequently reversed the TNF-α induced activation of transcription factor nuclear factor kappa B (NF-κB) signals and Endoplasmic Reticulum (ER) stress related apoptosis in chondrocytes. In addition, the protective effect of Sal and its inhibitory effect on PCAF as well as inflammatory and ER stress related markers were also found in mouse DMM model. Our studies have uncovered that aberrant PCAF accumulation contributes to the OA development and the mechanism underlying the anti-inflammatory and anti-apoptotic effect of Sal is in relation to the cross-talk between PCAF and NF-κB and ER stress cascades. Funding Statement: Financial supports from National Natural Science Foundation of China (NO. 81601983, NO.81501869). Declaration of Interests: The authors report no conflicts of interest. Ethics Approval Statement: The experimental protocols, animal use and postoperative animal care procedures were performed in line with the guidelines and principles of the Animal Care and Use Committee of Wenzhou Medical University (ethic code: wydw2014-0129). The whole study was approved by the medical ethics committee of The Second Affiliated Hospital of Wenzhou Medical University (number: LCKY-2017-36) and in accordance with the guidelines of the Declaration of Helsinki. Informed consent was obtained from every participant.

Abstract 1196: Induction of NKG2D ligand expression by CD8+ T cells through a nuclear factor-kappa B and P300/CBP-associated factor-dependent mechanism

Abstract The ligands for the natural-killer group 2 (NKG2D) protein are frequently expressed on tumor cell surfaces, rendering tumor cells susceptible to NKG2D-dependent immune cell attack. However, cancer cells escape from immune surveillance by downregulating NKG2D ligands. We previously discovered that the engagement of activated CD8+ T cells and tumor cells induces NKG2D ligands on tumor cells, but the underlying mechanism remains to be defined. TCGA database analyses suggested that the expression of CD40 and CD137L (their cognate receptors are CD40L and CD137, respectively, expressed on CD8+T cells) is associated with pan-NKG2D ligand expression in sarcoma and colorectal adenocarcinoma patients (Pearson’s correlation coefficient r=0.2225 and 0.1947, respectively; and P=.0003 and .0002, respectively). In agreement with these human data analysis results, both in vivo mouse tumor models and in vitro cell assays validated that upon the engagement with the cognate receptors, the tumor necrosis factor receptor superfamily (TNFRSF), particularly CD137 ligand and CD40, initiates the activation of nuclear factor-kappa B (NF-κB) signaling in tumor cells even in the absence of CD8+ T cells. The same as tumor and CD8+ T contact-dependent NKG2D ligand induction, this CD137L/CD40-mediated signaling activation was associated with elevated levels of acetyltransferase P300/CBP-associated factor (PCAF), whereas inhibition of phosphorylated NF-κB abrogated PCAF induction. Although the stimulation of CD137L/CD40-mediated signaling by cognate receptors is vital for NKG2D ligand upregulation, the inflammatory cytokines, including interferon γ (IFNγ) and tumor necrosis factor α (TNFα) released from CD8+ T cells also facilitate NKG2D ligand-induced immune surveillance. These inflammatory cytokines not only trigger NF-κB signaling and PCAF activation, and they also boost T cell chemotaxis, recruit large numbers of CD8+T cells to tumor sites, and result in high yield expression of NKG2D ligands. Collectively, our results unveil a novel mechanism of NKG2D ligand upregulation involving reverse signaling of CD40 and CD137L on in tumor cells, which along with inflammatory cytokines IFNγ and TNFα, stimulate downstream NF-κB and PCAF activation. Understanding this mechanism may help to develop induced NKG2D ligand-dependent T-cell therapy against cancers. Citation Format: Jiemiao Hu, Xueqing Xia, Shulin Li. Induction of NKG2D ligand expression by CD8+ T cells through a nuclear factor-kappa B and P300/CBP-associated factor-dependent mechanism [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1196.

Caffeic Acid Phenethyl Ester Inhibits UV-Induced MMP-1 Expression by Targeting Histone Acetyltransferases in Human Skin.

Caffeic acid phenethyl ester (CAPE), a naturally occurring bioactive compound, displays anti-inflammatory, anti-carcinogenic, and anti-microbial effects. However, the effect of CAPE on skin photoaging is unknown. Herein, we investigated the inhibitory effect of CAPE against ultraviolet (UV) irradiation-mediated matrix metalloproteinase (MMP)-1 expression and its underlying molecular mechanism. CAPE treatment suppressed UV-induced MMP-1 levels in both human dermal fibroblasts (HDF) and human skin tissues. While CAPE did not display any significant effects against the upstream regulatory pathways of MMP-1, CAPE was capable of reversing UV-mediated epigenetic modifications. CAPE suppressed UV-induced acetyl-histone H3 (Lys9) as well as total lysine acetylation in HDF cells. Similarly, CAPE also attenuated UV-induced lysine acetylations in human skin tissues, suggesting that the CAPE-mediated epigenetic alterations can be recapitulated in ex vivo conditions. CAPE was found to attenuate UV-induced histone acetyltransferase (HAT) activity in HDF. Notably, CAPE was able to directly inhibit the activity of several HATs including p300, CREP-binding protein (CBP), and p300/CBP-associated factor (PCAF), further confirming that CAPE can function as an epigenetic modulator. Thus, our study suggests that CAPE maybe a promising agent for the prevention of skin photoaging via targeting HATs.

Epigenetic Alterations Induced by Photothrombotic Stroke in the Rat Cerebral Cortex: Deacetylation of Histone h3, Upregulation of Histone Deacetylases and Histone Acetyltransferases.

Ischemic penumbra that surrounds a stroke-induced infarction core is potentially salvageable; however, mechanisms of its formation are not well known. Covalent modifications of histones control chromatin conformation, gene expression and protein synthesis. To study epigenetic processes in ischemic penumbra, we used photothrombotic stroke (PTS), a stroke model in which laser irradiation of the rat brain cortex photosensitized by Rose Bengal induces local vessel occlusion. Immunoblotting and immunofluorescence microscopy showed decrease in acetylation of lysine 9 in histone H3 in penumbra at 1, 4 or 24 h after PTS. This was associated with upregulation of histone deacetylases HDAC1 and HDAC2, but not HDAC4, which did not localize in the nuclei. HDAC2 was found in cell nuclei, HDAC4 in the cytoplasm and HDAC1 both in nuclei and cytoplasm. Histone acetyltransferases HAT1 and PCAF (p300/CBP associated factor) that acetylated histone H3 synthesis were also upregulated, but lesser and later. PTS increased localization of HDAC2 and HAT1 in astroglia. Thus, the cell fate in PTS-induced penumbra is determined by the balance between opposite tendencies leading either to histone acetylation and stimulation of gene expression, or to deacetylation and suppression of transcriptional processes and protein biosynthesis. These epigenetic proteins may be the potential targets for anti-stroke therapy.

Epigenetically Down-Regulated Acetyltransferase PCAF Increases the Resistance of Colorectal Cancer to 5-Fluorouracil

Only 10%–20% of colorectal cancer (CRC) patients observe effective responses to 5-fluorouracil (5-FU) based chemo-treatment. We used real-time PCR array and Western blot analysis to examine the expression alteration of acetyltransferases and deacetylases in 5-FU resistant CRC cell lines as compared to their respective parental CRC cell lines. Unlike other acetyltransferases and deacetylases, we found that the expression of acetyltransferase P300/CBP-associated factor (PCAF) is consistently decreased in three 5-FU resistant CRC cell lines. Similarly, knockdown of PCAF in HCT116 CRC parental cell line also increases the resistance to 5-FU and attenuates 5-FU-induced apoptosis. Mechanistically, we demonstrated that increased binding of trimethylated histone H3K27 in the promoter region of PCAF attenuated its transcription in 5-FU resistant HCT116/5-FU cells. Decreased PCAF impairs the acetylation of p53 and attenuates the p53-dependent transcription of p21, which results in the increased cyclin D1 and phosphorylation of Retinoblastoma 1. Conversely, overexpression of PCAF in CRC cell lines increases p21 and their susceptibility to 5-FU in vitro and in vivo. However, knockdown of p21 abolishes the beneficial effects of PCAF overexpression on increasing the sensitivity of HCT116/5-FU cells to 5-FU. Also, the reduced intensity of PCAF immunostaining was observed in the precancerous lesion, and microarray data from the public database further demonstrated the association between PCAF down-regulation and poor survival outcome. Our data suggest that PCAF-mediated p53 acetylation is an essential regulatory mechanism for increasing the susceptibility of CRC to 5-FU.

Complementary Roles of GCN5 and PCAF in Foxp3+ T-Regulatory Cells.

Functions of the GCN5-related N-acetyltransferase (GNAT) family of histone/protein acetyltransferases (HATs) in Foxp3+ T-regulatory (Treg) cells are unexplored, despite the general importance of these enzymes in cell biology. We now show that two prototypical GNAT family members, GCN5 (general control nonrepressed-protein 5, lysine acetyltransferase (KAT)2a) and p300/CBP-associated factor (p300/CBP-associated factor (PCAF), Kat2b) contribute to Treg functions through partially distinct and partially overlapping mechanisms. Deletion of Gcn5 or PCAF did not affect Treg development or suppressive function in vitro, but did affect inducible Treg (iTreg) development, and in vivo, abrogated Treg-dependent allograft survival. Contrasting effects were seen upon targeting of each HAT in all T cells; mice lacking GCN5 showed prolonged allograft survival, suggesting this HAT might be a target for epigenetic therapy in allograft recipients, whereas transplants in mice lacking PCAF underwent acute allograft rejection. PCAF deletion also enhanced anti-tumor immunity in immunocompetent mice. Dual deletion of GCN5 and PCAF led to decreased Treg stability and numbers in peripheral lymphoid tissues, and mice succumbed to severe autoimmunity by 3–4 weeks of life. These data indicate that HATs of the GNAT family have contributions to Treg function that cannot be replaced by the functions of previously characterized Treg HATs (CBP, p300, and Tip60), and may be useful targets in immuno-oncology.

Discovery of Pyrrolo[3,2- d]pyrimidin-4-one Derivatives as a New Class of Potent and Cell-Active Inhibitors of P300/CBP-Associated Factor Bromodomain.

Herein, we report the discovery of a series of new P300/CBP-associated factor (PCAF) bromodomain (BRD) inhibitors, which were obtained through a hit discovery process and subsequent structure-based optimization and structure-activity relationship analyses toward a retrieved hit compound (12). Among these inhibitors, ( R, R)-36n is the most potent one with an IC50 of 7 nM in homogeneous time-resolved fluorescence assay and a KD of 78 nM in isothermal titration calorimetry assay. This compound also exhibited activity against GCN5 and FALZ, but weak or no activity against other 29 BRD proteins and 422 kinases, indicating considerable selectivity. X-ray cocrystal structure analysis revealed the molecular interaction mode and the precise stereochemistry required for bioactivity. Cellular activity, preliminary RNA-seq analysis, and pharmacokinetic properties were also examined for this compound. Collectively, this study provides a versatile tool molecule to explore molecular mechanisms of PCAF BRD regulation and also offers a new lead compound for drug discovery targeting PCAF.

Downregulation of the transcriptional co-activator PCAF inhibits the proliferation and migration of vascular smooth muscle cells and attenuates NF-κB-mediated inflammatory responses

P300/CBP-associated factor (PCAF) regulates vascular inflammation. This study was to explore the effect of PCAF on the proliferation and migrationof vascular smooth muscle cells (VSMCs) and neointimal hyperplasia in balloon-injured rat carotid artery. Downregulation of PCAF remarkably suppressed VSMCs proliferation and migration induced by lipopolysaccharide, and also significantly inhibit the nuclear translocation of nuclear factor-kappaB p65. Meanwhile, downregulation of PCAF inhibited the mRNA expression of tumor necrosis factor-α and interleukin-6, and also the levels in culture supernatants. Moreover, downregulation of PCAF profoundly reduced the intima area and the ratio of intima area to media area in balloon-injured rat carotid artery. In addition, the expression of PCNA and NF-κB p65 in intima were decreased by downregulation of PCAF. These results highlight that PCAF may be a potential target for prevention and treatment of neointimal hyperplasia and restenosis after angioplasty.

Inhibition of p300/CBP-Associated Factor Attenuates Renal Tubulointerstitial Fibrosis through Modulation of NF-kB and Nrf2.

p300/CBP-associated factor (PCAF), a histone acetyltransferase, is involved in many cellular processes such as differentiation, proliferation, apoptosis, and reaction to cell damage by modulating the activities of several genes and proteins through the acetylation of either the histones or transcription factors. Here, we examined a pathogenic role of PCAF and its potential as a novel therapeutic target in the progression of renal tubulointerstitial fibrosis induced by non-diabetic unilateral ureteral obstruction (UUO) in male C57BL/6 mice. Administration of garcinol, a PCAF inhibitor, reversed a UUO-induced increase in the renal expression of total PCAF and histone 3 lysine 9 acetylation and reduced positive areas of trichrome and α-smooth muscle actin and collagen content. Treatment with garcinol also decreased mRNA levels of transforming growth factor-β, matrix metalloproteinase (MMP)-2, MMP-9, and fibronectin. Furthermore, garcinol suppressed nuclear factor-κB (NF-κB) and pro-inflammatory cytokines such as tumor necrosis factor-α and IL-6, whereas it preserved the nuclear expression of nuclear factor erythroid-derived 2-like factor 2 (Nrf2) and levels of Nrf2-dependent antioxidants including heme oxygense-1, catalase, superoxide dismutase 1, and NAD(P)H:quinone oxidoreductase 1. These results suggest that the inhibition of inordinately enhanced PCAF could mitigate renal fibrosis by redressing aberrant balance between inflammatory signaling and antioxidant response through the modulation of NF-κB and Nrf2.

P300/CBP-associated factor promotes autophagic degradation of \u03b4-catenin through acetylation and decreases prostate cancer tumorigenicity

δ-Catenin shares common binding partners with β-catenin. As acetylation and deacetylation regulate β-catenin stability, we searched for histone acetyltransferases (HATs) or histone deacetylases (HDACs) affecting δ-catenin acetylation status and protein levels. We showed that p300/CBP-associated factor (PCAF) directly bound to and acetylated δ-catenin, whereas several class I and class II HDACs reversed this effect. Unlike β-catenin, δ-catenin was downregulated by PCAF-mediated acetylation and upregulated by HDAC-mediated deacetylation. The HDAC inhibitor trichostatin A attenuated HDAC1-mediated δ-catenin upregulation, whereas HAT or autophagy inhibitors, but not proteasome inhibitors, abolished PCAF-mediated δ-catenin downregulation. The results suggested that PCAF-mediated δ-catenin acetylation promotes its autophagic degradation in an Atg5/LC3-dependent manner. Deletions or point mutations identified several lysine residues in different δ-catenin domains involved in PCAF-mediated δ-catenin downregulation. PCAF overexpression in prostate cancer cells markedly reduced δ-catenin levels and suppressed cell growth and motility. PCAF-mediated δ-catenin downregulation inhibited E-cadherin processing and decreased the nuclear distribution of β-catenin, resulting in the suppression of β-catenin/LEF-1-mediated downstream effectors. These data demonstrate that PCAF downregulates δ-catenin by promoting its autophagic degradation and suppresses δ-catenin-mediated oncogenic signals.

SUMOylation of G9a regulates its function as an activator of myoblast proliferation

The lysine methyltransferase G9a plays a role in many cellular processes. It is a potent repressor of gene expression, a function attributed to its ability to methylate histone and non-histone proteins. Paradoxically, in some instances, G9a can activate gene expression. However, regulators of G9a expression and activity are poorly understood. In this study, we report that endogenous G9a is SUMOylated in proliferating skeletal myoblasts. There are four potential SUMOylation consensus motifs in G9a. Mutation of all four acceptor lysine residues [K79, K152, K256, and K799] inhibits SUMOylation. Interestingly, SUMOylation does not impact G9a-mediated repression of MyoD transcriptional activity or myogenic differentiation. In contrast, SUMO-defective G9a is unable to enhance proliferation of myoblasts. Using complementation experiments, we show that the proliferation defect of primary myoblasts from conditional G9a-deficient mice is rescued by re-expression of wild-type, but not SUMOylation-defective, G9a. Mechanistically, SUMOylation acts as signal for PCAF (P300/CBP-associated factor) recruitment at E2F1-target genes. This results in increased histone H3 lysine 9 acetylation marks at E2F1-target gene promoters that are required for S-phase progression. Our studies provide evidence by which SUMO modification of G9a influences the chromatin environment to impact cell cycle progression.

Anacardic acid attenuates pressure-overload cardiac hypertrophy through inhibiting histone acetylases.

Cardiac hypertrophy has become a major cardiovascular problem wordwide and is considered the early stage of heart failure. Treatment and prevention strategies are needed due to the suboptimal efficacy of current treatment methods. Recently, many studies have demonstrated the important role of histone acetylation in myocardium remodelling along with cardiac hypertrophy. A Chinese herbal extract containing anacardic acid (AA) is known to possess strong histone acetylation inhibitory effects. In previous studies, we demonstrated that AA could reverse alcohol‐induced cardiac hypertrophy in an animal model at the foetal stage. Here, we investigated whether AA could attenuate cardiac hypertrophy through the modulation of histone acetylation and explored its potential mechanisms in the hearts of transverse aortic constriction (TAC) mice. This study showed that AA attenuated hyperacetylation of acetylated lysine 9 on histone H3 (H3K9ac) by inhibiting the expression of p300 and p300/CBP‐associated factor (PCAF) in TAC mice. Moreover, AA normalized the transcriptional activity of the heart nuclear transcription factor MEF2A. The high expression of cardiac hypertrophy‐linked genes (ANP, β‐MHC) was reversed through AA treatment in the hearts of TAC mice. Additionally, we found that AA improved cardiac function and survival rate in TAC mice. The current results further highlight the mechanism by which histone acetylation is controlled by AA treatment, which may help prevent and treat hypertrophic cardiomyopathy.

ALT cancer cells are specifically sensitive to lysine acetyl transferase inhibition.

Some cancer cells elongate their telomeres through the ALT (alternative lengthening of telomeres) pathway, which is based on homologous recombination for the addition of telomere repeats without telomerase activity. General control non-derepressible 5 (GCN5) and P300/CBP-associated factor (PCAF), two homologous lysine acetyltransferases, exert opposite effects on the ALT pathway, inhibiting or favoring it respectively. Here we show that ALT cells are particularly sensitive to the inhibition of acetyltransferases activities using Anacardic Acid (AA). AA treatment recapitulates the effect of PCAF knockdown on several ALT features, suggesting that AA decreased the ALT mechanism through the inhibition of lysine transferase activity of PCAF, but not that of GCN5. Furthermore, AA specifically sensitizes human ALT cells to radiation as compared to telomerase-positive cells suggesting that the inhibition of lysine acetyltransferases activity may be used to increase the radiotherapy efficiency against ALT cancers.