RESUMO
Aggregatibacter actinomycetemcomitans, a Gram-negative bacterium, is the causative agent of localized aggressive periodontitis. Attachment to a biotic surface is a critical first step in the A. actinomycetemcomitans infection process for which exopolysaccharides have been shown to be essential. In addition, the pga operon, containing genes encoding for biosynthetic proteins for poly-N-acetyl glucosamine (PNAG), plays a key role in A. actinomycetemcomitans virulence, as a mutant strain lacking the pga operon induces significantly less bone resorption. Among the genes in the pga operon, pgaB codes for a de-N-acetylase that is responsible for the deacetylation of the PNAG exopolysaccharide. Here we report the role of PgaB in regulation of virulence genes using a markerless, scarless deletion mutant targeting the coding region of the N-terminal catalytic domain of PgaB. The results demonstrate that the N-terminal, catalytic domain of PgaB is crucial for exopolysaccharide export.
Assuntos
Acetilesterase/genética , Acetilesterase/fisiologia , Aggregatibacter actinomycetemcomitans/enzimologia , Aggregatibacter actinomycetemcomitans/genética , Aggregatibacter actinomycetemcomitans/metabolismo , Genes Bacterianos/genética , Acetilglucosamina/imunologia , Acetilglucosamina/metabolismo , Aggregatibacter actinomycetemcomitans/patogenicidade , Proteínas de Bactérias/genética , Proteínas de Bactérias/fisiologia , Biofilmes/crescimento & desenvolvimento , Domínio Catalítico , Perfilação da Expressão Gênica , Regulação Bacteriana da Expressão Gênica , Fases de Leitura Aberta/genética , Óperon , Periodontite , Polissacarídeos Bacterianos , Deleção de Sequência , Virulência/genéticaRESUMO
The death of cells harboring defects in two distinct pathways implicates these pathways in the control of an essential process. Here we report that cells lacking OmpR and harboring constitutively active MalT undergo premature death that involves increased expression of the outer membrane porin LamB.
Assuntos
Proteínas de Bactérias/fisiologia , Proteínas de Ligação a DNA/fisiologia , Proteínas de Escherichia coli/fisiologia , Escherichia coli/crescimento & desenvolvimento , Escherichia coli/genética , Transativadores/fisiologia , Fatores de Transcrição/fisiologia , Transportadores de Cassetes de Ligação de ATP/genética , Transportadores de Cassetes de Ligação de ATP/fisiologia , Acetilesterase/genética , Acetilesterase/fisiologia , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/fisiologia , Proteínas de Bactérias/genética , Cistationina gama-Liase/genética , Cistationina gama-Liase/fisiologia , Proteínas de Ligação a DNA/genética , Proteínas de Escherichia coli/genética , Mutação , Porinas/genética , Porinas/fisiologia , Receptores Virais/genética , Receptores Virais/fisiologia , Proteínas Repressoras/genética , Proteínas Repressoras/fisiologia , Transativadores/genética , Fatores de Transcrição/genéticaRESUMO
Eukaryotic GCN5 acetyltransferases influence diverse biological processes by acetylating histones and non-histone proteins and regulating chromatin and gene-specific transcription as part of multiprotein complexes. In lower eukaryotes and invertebrates, these complexes include the yeast ADA complex that is still incompletely understood; the SAGA (Spt-Ada-Gcn5 acetylase) complexes from yeast to Drosophila that are mostly coactivators; and the ATAC (Ada Two-A containing) complex, only known in Drosophila and still poorly characterized. In contrast, vertebrate organisms, express two paralogous GCN5-like acetyltransferases (GCN5 and PCAF), which have been found so far only in SAGA-type complexes referred to hereafter as the STAGA (SPT3-TAF9-GCN5/PCAF acetylase) complexes. We now report the purification and characterization of vertebrate (human) ATAC-type complexes and identify novel components of STAGA. We show that human ATAC complexes incorporate in addition to GCN5 or PCAF (GCN5/PCAF), other epigenetic coregulators (ADA2-A, ADA3, STAF36, and WDR5), cofactors of chromatin assembly/remodeling and DNA replication machineries (POLE3/CHRAC17 and POLE4), the stress- and TGFbeta-activated protein kinase (TAK1/MAP3K7) and MAP3-kinase regulator (MBIP), additional cofactors of unknown function, and a novel YEATS2-NC2beta histone fold module that interacts with the TATA-binding protein (TBP) and negatively regulates transcription when recruited to a promoter. We further identify the p38 kinase-interacting protein (p38IP/FAM48A) as a novel component of STAGA with distant similarity to yeast Spt20. These results suggest that vertebrate ATAC-type and STAGA-type complexes link specific extracellular signals to modification of chromatin structure and regulation of the basal transcription machinery.
Assuntos
Acetilesterase/fisiologia , Histona Acetiltransferases/química , Histonas/química , Fosfoproteínas/metabolismo , Proteínas de Saccharomyces cerevisiae/química , Proteína de Ligação a TATA-Box/química , Fatores de Transcrição/metabolismo , Fatores de Transcrição de p300-CBP/química , Acetilesterase/química , Animais , Drosophila , Genes Reporter , Humanos , Fosfoproteínas/fisiologia , Dobramento de Proteína , Estrutura Terciária de Proteína , Proteína de Ligação a TATA-Box/metabolismo , Fatores de Transcrição/fisiologia , Transcrição Gênica , Técnicas do Sistema de Duplo-Híbrido , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
Influenza C virus uses 9-O-acetyl-N-acetylaneuraminic acid (9-O-acetyl-Neu5Ac) as a receptor determinant for attachment to cells. The virus contains an acetylesterase which releases acetyl residues from position C-9 of sialic acid thereby inactivating the receptors. A synthetic sialic acid analogue, 9-N-acetyl-Neu5Ac, was attached to cell surface glycoconjugates by purified sialyltransferase and analyzed for its ability to substitute the 9-O-acetylated sialic acid. Erythrocytes which have been modified to contain either 9-O-acetyl-Neu5Ac or 9-N-acetyl-Neu5Ac were agglutinated by influenza C virus to the same titer. However, in contrast to the 9-O-acetyl group the 9-N-acetyl residue is resistant to cleavage by the viral acetylesterase. This characteristic property (recognition as a receptor determinant by influenza C virus, but resistance against the action of the receptor-destroying enzyme) makes this synthetic analogue a valuable tool to analyze the role of the receptor-destroying enzyme for an influenza C virus infection.
Assuntos
Acetilesterase/fisiologia , Eritrócitos/metabolismo , Gammainfluenzavirus/metabolismo , Receptores Virais/efeitos dos fármacos , Ácidos Siálicos , Animais , Galinhas , Agregação Eritrocítica , Eritrócitos/ultraestrutura , Glicoproteínas/metabolismo , Gammainfluenzavirus/fisiologia , Infecções por Orthomyxoviridae/sangue , Infecções por Orthomyxoviridae/etiologia , Infecções por Orthomyxoviridae/fisiopatologia , Receptores Virais/metabolismo , Ácidos Siálicos/metabolismo , Sialiltransferases/metabolismo , Sialiltransferases/fisiologia , beta-D-Galactosídeo alfa 2-6-SialiltransferaseRESUMO
The nature of the receptor-destroying enzyme (RDE) of influenza C virus has been elucidated by analyzing its effect on the haemagglutination inhibitors rat alpha 1-macroglobulin (RMG) and bovine submandibulary mucin (BSM), respectively. The inhibitory activity of both compounds is abolished by incubation with influenza C virus. After inactivation, RMG and BSM were found to contain reduced amounts of N-acetyl-9-O-acetylneuraminic acid (Neu5,9Ac2) and increased amounts of N-acetylneuraminic acid (Neu5Ac). H.p.l.c. analysis revealed that purified Neu5,9Ac2 is converted to Neu5Ac by incubation with influenza C virus. These results demonstrate that RDE of influenza C virus is neuraminate-O-acetylesterase [N-acyl-9(4)-O-acetylneuraminate O-acetylhydrolase (EC 3.1.1.53)]. The data also indicate that haemagglutination-inhibition (HI) by RMG and BSM and most likely virus attachment to cell surfaces involves binding of influenza C virus to Neu5,9Ac2.