RESUMEN
The COQ2 gene in Saccharomyces cerevisiae encodes a Coq2 (p-hydroxybenzoate:polyprenyl transferase), which is required in the biosynthetic pathway of CoQ (ubiquinone). This enzyme catalyses the prenylation of p-hydroxybenzoate with an all-trans polyprenyl group. We have isolated cDNA which we believe encodes the human homologue of COQ2 from a human muscle and liver cDNA library. The clone contained an open reading frame of length 1263 bp, which encodes a polypeptide that has sequence homology with the Coq2 homologues in yeast, bacteria and mammals. The human COQ2 gene, when expressed in yeast Coq2 null mutant cells, rescued the growth of this yeast strain in the absence of a non-fermentable carbon source and restored CoQ biosynthesis. However, the rate of CoQ biosynthesis in the rescued cells was lower when compared with that in cells rescued with the yeast COQ2 gene. CoQ formed when cells were incubated with labelled decaprenyl pyrophosphate and nonaprenyl pyrophosphate, showing that the human enzyme is active and that it participates in the biosynthesis of CoQ.
Asunto(s)
Transferasas Alquil y Aril/genética , Regulación Enzimológica de la Expresión Génica/genética , Ubiquinona/biosíntesis , Transferasas Alquil y Aril/deficiencia , Transferasas Alquil y Aril/metabolismo , Secuencia de Aminoácidos/genética , Secuencia de Bases/genética , Clonación Molecular/métodos , Prueba de Complementación Genética/métodos , Humanos , Hígado/química , Hígado/metabolismo , Proteínas Mitocondriales/genética , Datos de Secuencia Molecular , Músculo Esquelético/química , Músculo Esquelético/metabolismo , Mutación/genética , Especificidad de Órganos/genética , Valor Predictivo de las Pruebas , Señales de Clasificación de Proteína/genética , ARN Mensajero/genética , Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/enzimología , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/crecimiento & desarrollo , Proteínas de Saccharomyces cerevisiae/genética , Análisis de Secuencia de ADN/métodosRESUMEN
Tanis is a recently described protein reported to be a putative receptor for serum amyloid A and found to be dysregulated with diabetes in the Israeli sand rat Psamommys obesus. Tanis has also been identified as a selenoprotein, one of the first two identified membrane selenoproteins. We determined mRNA expression of the human homologue of Tanis, SelS/AD-015, in skeletal muscle and adipose tissue biopsies obtained from 10 type 2 diabetic patients and 11 age- and weight-matched healthy subjects. Expression of Tanis/SelS mRNA in skeletal muscle and adipose tissue biopsies was similar between diabetic and control subjects. A subset of subjects underwent a euglycemic-hyperinsulinemic clamp, and adipose tissue expression of Tanis/SelS was determined after in vivo insulin stimulation. Adipose tissue Tanis/SelS mRNA expression was unchanged after insulin infusion in control subjects, whereas Tanis/SelS mRNA increased in seven of eight subjects following insulin stimulation in diabetic subjects. Skeletal muscle and adipose tissue Tanis/SelS mRNA expression were positively correlated with plasma serum amyloid A. In conclusion, there is a strong trend toward upregulation of Tanis/SelS following insulin infusion in adipose tissue from type 2 diabetic subjects. Moreover, the positive relationship between Tanis mRNA and the acute-phase protein serum amyloid A suggests an interaction between innate immune system responses and Tanis expression in muscle and adipose tissue.
Asunto(s)
Tejido Adiposo/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Proteínas de la Membrana/metabolismo , Músculo Esquelético/metabolismo , Proteína Amiloide A Sérica/metabolismo , Estudios de Casos y Controles , Técnica de Clampeo de la Glucosa , Humanos , Insulina/farmacología , Masculino , Proteínas de la Membrana/genética , Persona de Mediana Edad , ARN Mensajero/metabolismo , Selenoproteínas , Regulación hacia ArribaRESUMEN
The gene of the p85alpha regulatory subunit of phosphatidylinositol (PI) 3-kinase gives rise to several splice variants. We hypothesized that the expression of p85alpha splice variants may be altered in skeletal muscle from subjects with type 2 diabetes mellitus. Skeletal muscle biopsies were obtained from nine type 2 diabetic and eight healthy men, matched for age, body mass index (BMI) and physical fitness. PI 3-kinase activity in skeletal muscle following in vitro insulin stimulation was reduced in subjects with type 2 diabetes. p85alpha mRNA was elevated fourfold in type 2 diabetic as compared to healthy control subjects ( P<0.05). p85alpha mRNA abundance was positively correlated with plasma insulin concentration ( P<0.01) and serum glucose concentration ( P<0.01). Despite this, protein levels of p85alpha, p55alpha, and the novel human p50alpha were not altered in type 2 diabetic subjects. Thus, although gene expression of full-length p85alpha is increased in skeletal muscle from type 2 diabetics, this is not reflected by increased protein levels. Therefore, defects in PI 3-kinase activity are likely due to impaired activation of the enzyme rather than changes in protein expression of the isoforms of the regulatory subunit.
