RESUMO
Sialic acids (Sias), 9-carbon-backbone sugars, are among the most complex and versatile molecules of life. As terminal residues of glycans on proteins and lipids, Sias are key elements of glycotopes of both cellular and microbial lectins and thus act as important molecular tags in cell recognition and signaling events. Their functions in such interactions can be regulated by post-synthetic modifications, the most common of which is differential Sia-O-acetylation (O-Ac-Sias). The biology of O-Ac-Sias remains mostly unexplored, largely because of limitations associated with their specific in situ detection. Here, we show that dual-function hemagglutinin-esterase envelope proteins of nidoviruses distinguish between a variety of closely related O-Ac-Sias. By using soluble forms of hemagglutinin-esterases as lectins and sialate-O-acetylesterases, we demonstrate differential expression of distinct O-Ac-sialoglycan populations in an organ-, tissue- and cell-specific fashion. Our findings indicate that programmed Sia-O-acetylation/de-O-acetylation may be critical to key aspects of cell development, homeostasis, and/or function.
Assuntos
Acetilesterase/biossíntese , Hemaglutininas Virais/genética , Ácido N-Acetilneuramínico/genética , Ácidos Siálicos/genética , Proteínas Virais de Fusão/genética , Acetilação , Acetilesterase/genética , Animais , Regulação da Expressão Gênica , Genoma , Hemaglutininas Virais/química , Hemaglutininas Virais/metabolismo , Humanos , Lipídeos/química , Lipídeos/genética , Mamíferos , Ácido N-Acetilneuramínico/química , Ácido N-Acetilneuramínico/metabolismo , Nidovirales/química , Proteínas/química , Proteínas/genética , Ácidos Siálicos/química , Especificidade da Espécie , Proteínas Virais de Fusão/química , Proteínas Virais de Fusão/metabolismoRESUMO
The placenta plays an important role as a regulator of fetal nutrition and growth throughout development and placental factors contribute to gestational abnormalities such as preeclampsia. This study describes the genome-wide gene expression profiles of a large (n = 60) set of human placentas in order to uncover gene expression patterns associated with preeclampsia. In addition to confirming changes in expression of soluble factors associated with preeclampsia such as sFLT1 (soluble fms-like tyrosine kinase-1), sENG (soluble endoglin), and INHA (inhibin alpha), we also find changes in immune-associated signaling pathways, offering a potential upstream explanation for the shallow trophoblast invasion and inadequate uterine remodeling typically observed in pathogenesis of preeclampsia. Notably, we also find evidence of preeclampsia-associated placental upregulation of sialic acid acetylesterase (SIAE), a gene functionally associated with autoimmune diseases.
Assuntos
Acetilesterase/biossíntese , Pré-Eclâmpsia/metabolismo , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/biossíntese , Antígenos CD/biossíntese , Endoglina , Feminino , Estudo de Associação Genômica Ampla , Humanos , Recém-Nascido , Inibinas/biossíntese , Masculino , Pré-Eclâmpsia/etiologia , Gravidez , Análise Serial de Proteínas , Receptores de Superfície Celular/biossíntese , Trofoblastos/fisiologia , Regulação para CimaRESUMO
Acyl analogs of PAF are the major products synthesized during agonist stimulation of endothelial cells. We have previously shown that PAF: 1-acyl-2-lyso-sn-glycero-3-phosphocholine transacetylase in calf pulmonary artery endothelial cells is activated by ATP through protein phosphorylation, and the increase in transacetylase activity by ATP contributes to the biosynthesis of acyl analogs of PAF (J. Biol. Chem. 272, 17431-17437, 1997). To understand the mechanisms(s) by which ATP stimulates acyl analogs of PAF production, we have identified the subtypes of the purinergic receptor that are linked to the activation of two enzymes involved in the generation of acyl analogs of PAF, namely, transacetylase and phospholipase A2. Experiments with transient transfection of the cells with antisense and sense thio-oligonucleotide to cytosolic phospholipase A2 (cPLA2) were also performed to evaluate whether downstream activation of cPLA2 is involved in ATP-receptor mediated induction of arachidonate release and synthesis of radylacetyl-GPC. We found that the P2u/P2Y2 receptor, which recognizes a pyrimidine nucleotide, UTP, as well as purine nucleotides, shows a potency profile of UTP > ATP = ATP gamma S > 2-methylthio-ATP in mediating the activation of PAF: lysophospholipid transacetylase. On the other hand, ADP beta S and 2-methylthio-ATP have similar potencies as ATP but have lower potencies than UTP and ATP gamma S in stimulating the release of arachidonate. These results suggest that both P2u/P2Y2 and P2y/P2Y1 receptor subtypes promote arachidonate release. In addition, transient transfection of endothelial cells with cPLA2 antisense but not the sense thio-oligonucleotide inhibited the stimulation of arachidonate release and [3H]acetate incorporation into radyl[3H]acetyl-GPC. Thus, our data suggest that a receptor-mediated process is involved in the activation of transacetylase for the induced synthesis of acyl analogs of PAF in endothelial cells. Furthermore, it is likely that cPLA2 supplies the lysophospholipids as substrates for the transacetylation reaction.
Assuntos
Acetilesterase/biossíntese , Ácido Araquidônico/metabolismo , Endotélio Vascular/metabolismo , Fosfolipases A/fisiologia , Fator de Ativação de Plaquetas/fisiologia , Receptores Purinérgicos/fisiologia , Trifosfato de Adenosina/análogos & derivados , Trifosfato de Adenosina/farmacologia , Animais , Ácido Araquidônico/farmacologia , Bovinos , Linhagem Celular , Relação Dose-Resposta a Droga , Lisofosfolipídeos/biossíntese , Oligonucleotídeos/farmacologia , Fosfolipases A2 , Tionucleotídeos/farmacologia , Transfecção , Uridina Trifosfato/farmacologiaAssuntos
Acetilesterase , Técnicas Biossensoriais , Dependência de Heroína/diagnóstico , Heroína/análise , Proteínas Recombinantes , Acetilesterase/biossíntese , Acetilesterase/química , Sequência de Aminoácidos , Animais , Bovinos , Clonagem Molecular/métodos , Colorimetria/métodos , Sequência Consenso , DNA Recombinante , Esterases/química , Heroína/metabolismo , Dependência de Heroína/epidemiologia , Humanos , Camundongos , Dados de Sequência Molecular , Moraxella/enzimologia , Morfina/análise , Derivados da Morfina/análise , Coelhos , Ratos , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Rhodococcus/metabolismo , Sensibilidade e Especificidade , Homologia de Sequência de Aminoácidos , Suínos , Reino Unido/epidemiologiaRESUMO
The hemagglutinin-esterase (HE) protein of mouse hepatitis virus (MHV) is an optional envelope protein present in only some MHV isolates. Its expression is regulated by the copy number of a UCUAA pentanucleotide sequence present in the leader sequence of the viral genomic RNA. The functional significance of this viral protein so far is not clear. In this report, we compared the neuropathogenicity of two MHV isolates, JHM(2) and JHM(3), which express different amounts of HE protein. Intracerebral inoculation of these two viruses into C57BL/6 mice showed that JHM(2), which expresses an abundant amount of HE protein, was more neurovirulent than JHM(3), which expresses very little HE. Histopathology showed that early in infection, JHM(2) infected primarily neurons, while JHM(3) infected mainly glial cells. JHM(3) eventually infected neurons and caused a delayed death relative to JHM(2)-infected mice, suggesting that the progression of JHM(3) infection in the central nervous system was slower than JHM(2). In vitro infection of JHM(3) in primary mixed glial cell cultures of astrocyte-enriched cultures yielded higher virus titers than JHM(2), mimicking the preferential growth of JHM(3) in glial cells in vivo. These findings suggest that the reduced neuropathogenicity of JHM(3) may correlate with its preferential growth in glial cells. Sequence analysis showed that the S genes of these two viruses are identical, thus ruling out the S gene as the cause of the difference in neuropathogenicity between these two viruses. We conclude that the HE protein contributes to viral neuropathogenicity by influencing either the rate of virus spread, viral cell tropism or both.