RESUMEN
A systematic review and a meta-analysis were performed to quantify the accumulated information from genetic association studies investigating the impact of the CYP4F2 rs2108622 (p.V433M) polymorphism on coumarin dose requirement. An additional aim was to explore the contribution of the CYP4F2 variant in comparison with, as well as after stratification for, the VKORC1 and CYP2C9 variants. Thirty studies involving 9,470 participants met prespecified inclusion criteria. As compared with CC-homozygotes, T-allele carriers required an 8.3% (95% confidence interval (CI): 5.6-11.1%; P < 0.0001) higher mean daily coumarin dose than CC homozygotes to reach a stable international normalized ratio (INR). There was no evidence of publication bias. Heterogeneity among studies was present (I(2) = 43%). Our results show that the CYP4F2 p.V433M polymorphism is associated with interindividual variability in response to coumarin drugs, but with a low effect size that is confirmed to be lower than those contributed by VKORC1 and CYP2C9 polymorphisms.
Asunto(s)
Cumarinas/administración & dosificación , Sistema Enzimático del Citocromo P-450/genética , Polimorfismo Genético/genética , Anciano , Anciano de 80 o más Años , Algoritmos , Alelos , Hidrocarburo de Aril Hidroxilasas/genética , Estudios de Cohortes , Cumarinas/uso terapéutico , Estudios Transversales , Citocromo P-450 CYP2C9 , Familia 4 del Citocromo P450 , Etnicidad , Humanos , Relación Normalizada Internacional , Persona de Mediana Edad , Oxigenasas de Función Mixta/genética , Sesgo de Publicación , Factores Sexuales , Vitamina K Epóxido ReductasasRESUMEN
Selective disintegration of membrane-enclosed autophagic bodies is a feature of eukaryotic cells not studied in detail. Using a Saccharomyces cerevisiae mutant defective in autophagic-body breakdown, we identified and characterized Aut5p, a glycosylated integral membrane protein. Site-directed mutagenesis demonstrated the relevance of its putative lipase active-site motif for autophagic-body breakdown. aut5Delta cells show reduced protein turnover during starvation and are defective in maturation of proaminopeptidase I. Most recently, by means of the latter phenotype, Aut5p was independently identified as Cvt17p. In this study we additionally checked for effects on vacuolar acidification and detected mature vacuolar proteases, both of which are prerequisites for autophagic-body lysis. Furthermore, biologically active hemagglutinin-tagged Aut5p (Aut5-Ha) localizes to the endoplasmic reticulum (nuclear envelope) and is targeted to the vacuolar lumen independent of autophagy. In pep4Delta cells immunogold electron microscopy located Aut5-Ha at approximately 50-nm-diameter intravacuolar vesicles. Characteristic missorting in vps class E and fab1Delta cells, which affects the multivesicular body (MVB) pathway, suggests vacuolar targeting of Aut5-Ha similar to that of the MVB pathway. In agreement with localization of Aut5-Ha at intravacuolar vesicles in pep4Delta cells and the lack of vacuolar Aut5-Ha in wild-type cells, our pulse-chase experiments clearly indicated that Aut5-Ha degradation with 50 to 70 min of half-life is dependent on vacuolar proteinase A.
Asunto(s)
Autofagia , Hidrolasas de Éster Carboxílico/metabolismo , Lipasa/metabolismo , Glicoproteínas de Membrana/metabolismo , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae/enzimología , Vacuolas/enzimología , Secuencias de Aminoácidos , Secuencia de Aminoácidos , Ácido Aspártico Endopeptidasas/metabolismo , Proteínas Relacionadas con la Autofagia , Sitios de Unión , Hidrolasas de Éster Carboxílico/genética , Hidrolasas de Éster Carboxílico/aislamiento & purificación , Genes Fúngicos , Glicoproteínas/metabolismo , Semivida , Lipasa/genética , Lipasa/aislamiento & purificación , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/aislamiento & purificación , Microscopía Inmunoelectrónica , Datos de Secuencia Molecular , Fosfotransferasas (Aceptor de Grupo Alcohol)/genética , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Transporte de Proteínas , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/ultraestructura , Homología de Secuencia de Aminoácido , Vacuolas/ultraestructuraRESUMEN
Autophagy is a degradative transport pathway that delivers cytosolic proteins to the lysosome (vacuole). Cytosolic proteins appear inside the vacuole enclosed in autophagic vesicles. These autophagic vesicles are broken down in the vacuole together with their cytosolic content. The breakdown of vesicular transport intermediates is a unique feature of autophagy. We here identify Aut4p, a component essential for the disintegration of autophagic vesicles, inside the vacuole of S. cerevisiae cells. Aut4p is a putative integral membrane protein with limited homologies to permeases. Chromosomal deletion of AUT4 has no obvious influence on growth, vacuolar acidification and the activities of vacuolar proteinases. Like proteinase B-deficient cells, aut4-deleted cells show a partial reduction in total protein breakdown during nitrogen starvation. A biologically active fusion protein of Aut4p and the green fluorescent protein is visualized at the vacuolar membrane and in punctate structures attached to the vacuole.