AAC(6a2)-Ib is an important aminoglycoside resistance enzyme to target with enzymatic inhibitors. An in silico screening approach was used to identify potential inhibitors from the ChemBridge ...library. Several compounds were identified, of which two of them, 4-(2-{1-(3-methylphenyl)-4,6-dioxo-2-thioxotetrahydro-5(2H)-pyri m idinylidenemethyl}phenoxy)methylbenzoic acid and 2-{5-(4,6-dioxo-1,3-diphenyl-2-thioxotetrahydro-5(2H)-pyrimidinyl i dene)methyl-2-furyl}benzoic acid, showed micromolar activity in inhibiting acetylation of kanamycin A. These compounds are predicted to bind the aminoglycoside binding site of AAC(6a2)-Ib and exhibited competitive inhibition against kanamycin A.
Histone deacetylase (HDAC) is an attractive target for cancer therapy because it plays a key role in gene expression and carcinogenesis. N-hydroxy-7-(2-naphthylthio) heptanomide (HNHA) is a novel ...synthetic HDAC inhibitor (HDACI) that shows better pharmacological properties than a known HDACI present in the human fibrosarcoma cell: suberoylanilide hydroxamic acid (SAHA). Here, we investigate the anti-cancer activity of HNHA against breast cancer both in vitro and in vivo. HNHA arrested the cell cycle at the G1/S phase via p21 induction, which led to profound inhibition of cancer cell growth in vitro. In addition, HNHA-treated cells showed markedly decreased levels of VEGF and HIF-1 alpha than SAHA and fumagillin (FUMA) when accompanied by increased histone acetylation. HNHA significantly inhibited tumor growth in an in vivo mouse xenograft model. HNHA-treated mice survived significantly longer than SAHA- and FUMA-treated mice. Dynamic MRI showed significantly decreased blood flow in the HNHA-treated mice, implying that HNHA inhibits tumor neovascularization. This finding was accompanied by marked reductions of proangiogenic factors and significant induction of angiogenesis inhibitors in tumor tissues. We have shown that HNHA is an effective anti-tumor agent in breast cancer cells in vitro and in breast cancer xenografts in vivo. Collectively, these findings indicate that HNHA may be a potent anti-cancer agent against breast cancer due to its multi-faceted inhibition of HDAC activity, as well as anti-angiogenesis activity. (Cancer Sci 2011; 102: 343-350)
Asthmatic airway smooth muscle (ASM) expresses interferon-g-inducible protein-10 (CXCL10), a chemokine known to mediate mast cell migration into ASM bundles that has been reported in the airways of ...asthmatic patients. CXCL10 is elevated in patients suffering from viral exacerbations of asthma and in patients with chronic obstructive pulmonary disease (COPD), diseases in which corticosteroids are largely ineffective. IFNg and TNFa synergistically induce CXCL10 release from human ASM cells in a steroid-insensitive manner, via an as yet undefined mechanism. We report that TNFa activates the classical NF-IordmB (n uclear f actor IordmB) pathway, whereas IFNg activates JAK2/STAT-1a and that inhibition of the JAK/STAT pathway is more effective in abrogating CXCL10 release than the steroid fluticasone. The synergy observed with TNFa and IFNg together, however, did not lie at the level of NF-IordmB activation, STAT-1a phosphorylation, or in vivo binding of these transcription factors to the CXCL10 promoter. Stimulation of human ASM cells with TNFa and IFNg induced histone H4 but not histone H3 acetylation at the CXCL10 promoter, although no synergism was observed when both cytokines were combined. We show, however, that TNFa and IFNg exert a synergistic effect on the recruitment of CREB-binding protein (CBP) to the CXCL10, which is accompanied by increased RNA polymerase II. Our results provide evidence that synergism between TNFa and IFNg lies at the level of coactivator recruitment in human ASM and suggest that inhibition of JAK/STAT signaling may be of therapeutic benefit in steroid-resistant airway disease.
Acetylation is a protein post-translational modification (PTM) that can affect a variety of cellular processes. In bacteria, two PTM
ε-acetylation mechanisms have been identified: ...non-enzymatic/chemical acetylation via acetyl phosphate or acetyl coenzyme A and enzymatic acetylation via protein acetyltransferases. Prior studies have shown that extensive acetylation of
ε-lysine residues of numerous proteins from a variety of bacteria occurs via non-enzymatic acetylation. In
, new
ε-lysine acetyltransferases (KATs) that enzymatically acetylate other proteins have been identified, thus expanding the repertoire of protein substrates that are potentially regulated by acetylation. Therefore, we designed a study to leverage the wealth of structural data in the Protein Data Bank (PDB) to determine: (1) the 3D location of lysine residues on substrate proteins that are acetylated by
KATs, and (2) investigate whether these residues are conserved on 3D structures of their homologs. Five
KAT substrate proteins that were previously identified as being acetylated by YiaC and had 3D structures in the PDB were selected for further analysis: adenylate kinase (Adk), isocitrate dehydrogenase (Icd), catalase HPII (KatE), methionyl-tRNA formyltransferase (Fmt), and a peroxide stress resistance protein (YaaA). We methodically compared over 350 protein structures of these
enzymes and their homologs; to accurately determine lysine residue conservation requires a strategy that incorporates both flexible structural alignments and visual inspection. Moreover, our results revealed discrepancies in conclusions about lysine residue conservation in homologs when examining linear amino acid sequences compared to 3D structures.
The Myc oncoprotein family comprises transcription factors that control multiple cellular functions and are widely involved in oncogenesis. Here we report the identification of Myc-nick, a ...cytoplasmic form of Myc generated by calpain-dependent proteolysis at lysine 298 of full-length Myc. Myc-nick retains conserved Myc box regions but lacks nuclear localization signals and the bHLHZ domain essential for heterodimerization with Max and DNA binding. Myc-nick induces a-tubulin acetylation and altered cell morphology by recruiting histone acetyltransferase GCN5 to microtubules. During muscle differentiation, while the levels of full-length Myc diminish, Myc-nick and acetylated a-tubulin levels are increased. Ectopic expression of Myc-nick accelerates myoblast fusion, triggers the expression of myogenic markers, and permits Myc-deficient fibroblasts to transdifferentiate in response to MyoD. We propose that the cleavage of Myc by calpain abrogates the transcriptional inhibition of differentiation by full-length Myc and generates Myc-nick, a driver of cytoplasmic reorganization and differentiation.
Although the pathophysiological processes involved in dopamine (DA) neuron degeneration in Parkinson's disease (PD) are not completely known, apoptotic cell death has been suggested to be involved ...and can be modeled in DAergic cell lines using the mitochondrial toxin 1-methyl-4-phenylpyridinium (MPP super(+)). Recently, it has been suggested that histone deacetylase inhibitors (HDACIs) may reduce apoptotic cell death in various model systems. However, their utility in interfering with ABABABcell death remains unclear. The HDACIs sodium butyrate (NaB), valproate (VPA) and suberoylanilide hydroxamic acid (SAHA) were tested for their ability to prevent MPP super(+)-mediated cytotoxicity in human derived SK-N-SH and rat derived MES 23.5 cells. All three HDACIs at least partially prevented MPP super(+)-mediated apoptotic cell death. The protective effects of these HDACIs coincided with significant increases in histone acetylation. These results suggest that HDACIs may be potentially neuroprotective against ABABABcell death and should be explored further in animal models of PD.
Chemokines play important roles in asthma. Prostaglandin I sub(2) (PGI sub(2)) analogue is recently suggested as a candidate for treating asthma. However, the effects of PGI sub(2) analogues on the ...expression of Th1- and Th2-related chemokines are unknown. To this end, we investigated the in vitro effects of PGI sub(2) analogues on the expression of Th1-related chemokine interferon- gamma -inducible protein-10 (IP-10/CXCL10) and Th2-related chemokine macrophage-derived chemokine (MDC/CCL22) in human monocytes. The human monocytes were pretreated with iloprost and treprostinil before lipopolysaccharide (LPS) stimulation. IP-10 and MDC were measured by ELISA. Intracellular signaling was investigated by cyclic adenosine monophosphate (cAMP) assay, western blot and chromatin immunoprecipitation. PGI sub(2) analogues enhanced MDC, but suppressed IP-10 expression in LPS-stimulated monocytes. These effects were reversed by the I prostanoid (IP) receptor antagonist (CAY10449), peroxisomal proliferators-activated receptor (PPAR)- alpha antagonist (GW6741) and PPAR- gamma antagonist (GW9662). PGI sub(2) analogues increased intracellular cAMP levels. Forskolin, an adenyl cyclase activator, conferred similar effects. PGI sub(2) analogue-enhanced MDC expression was reduced by nuclear factor (NF) Kappa B inhibitor (BAY 117085) and mitogen-activated protein kinase (MAPK)-p38 inhibitor (SB203580). PGI sub(2) analogues up-regulated phospho-p65 and phospho-p38 but down-regulated phospho-ERK expression. Iloprost enhanced H3 acetylation in MDC promoter area and suppressed H3 acetylation, H3K4, and H3K36 trimethylation in IP-10 promoter area. PGI sub(2) analogues enhanced MDC expression via the I prostanoid-receptor-cAMP, PPAR- alpha and PPAR- gamma , NF Kappa B-p65, MAPK-p38-ATF2 pathways and increasing histone acetylation, and suppressed IP-10 expression via the IP-receptor-cAMP, PPAR- gamma , MAPK-ERK-ELK1 pathways and inhibiting histone acetylation and trimethylation in LPS-stimulated monocytes. PGI sub(2) analogues may therefore increase Th2 recruitment and inflammation.
Heat shock protein 90 (Hsp90) is an ATP-dependent molecular chaperone which is essential in eukaryotes. It is required for the activation and stabilization of a wide variety of client proteins and ...many of them are involved in important cellular pathways. Since Hsp90 affects numerous physiological processes such as signal transduction, intracellular transport, and protein degradation, it became an interesting target for cancer therapy. Structurally, Hsp90 is a flexible dimeric protein composed of three different domains which adopt structurally distinct conformations. ATP binding triggers directionality in these conformational changes and leads to a more compact state. To achieve its function, Hsp90 works together with a large group of cofactors, termed co-chaperones. Co-chaperones form defined binary or ternary complexes with Hsp90, which facilitate the maturation of client proteins. In addition, posttranslational modifications of Hsp90, such as phosphorylation and acetylation, provide another level of regulation. They influence the conformational cycle, co-chaperone interaction, and inter-domain communications. In this review, we discuss the recent progress made in understanding the Hsp90 machinery.
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