title: Membrane-association of mRNA decapping factors is independent of stress in budding yeast.
abstract: Recent evidence has suggested that the degradation of mRNA occurs on translating ribosomes or alternatively within RNA granules called P bodies, which are aggregates whose core constituents are mRNA decay proteins and RNA. In this study, we examined the mRNA decapping proteins, Dcp1, Dcp2, and Dhh1, using subcellular fractionation. We found that decapping factors co-sediment in the polysome fraction of a sucrose gradient and do not alter their behaviour with stress, inhibition of translation or inhibition of the P body formation. Importantly, their localisation to the polysome fraction is independent of the RNA, suggesting that these factors may be constitutively localised to the polysome. Conversely, polysomal and post-polysomal sedimentation of the decapping proteins was abolished with the addition of a detergent, which shifts the factors to the non-translating RNP fraction and is consistent with membrane association. Using a membrane flotation assay, we observed the mRNA decapping factors in the lower density fractions at the buoyant density of membrane-associated proteins. These observations provide further evidence that mRNA decapping factors interact with subcellular membranes, and we suggest a model in which the mRNA decapping factors interact with membranes to facilitate regulation of mRNA degradation.
authors.affiliation: Department of Molecular Biology, Umeå University, SE-901 87 Umeå, Sweden.
authors.forename: Susanne
authors.initials: S
authors.lastname: Huch
authors.affiliation: Department of Molecular Biology, Umeå University, SE-901 87 Umeå, Sweden.
authors.forename: Jessie
authors.initials: J
authors.lastname: Gommlich
authors.affiliation: Department of Molecular Biology, Umeå University, SE-901 87 Umeå, Sweden.
authors.forename: Mridula
authors.initials: M
authors.lastname: Muppavarapu
authors.affiliation: Department of Urology, University of Rochester, Rochester, NY 14642, USA.
authors.forename: Carla
authors.initials: C
authors.lastname: Beckham
authors.affiliation: Department of Molecular Biology, Umeå University, SE-901 87 Umeå, Sweden.
authors.forename: Tracy
authors.initials: T
authors.lastname: Nissan
mesh_terms: D002453:Cell Cycle / Q000235:genetics; D002458:Cell Fractionation / Q000379:methods; D002462:Cell Membrane / Q000378:metabolism* / Q000648:ultrastructure; D002499:Centrifugation, Density Gradient; D053487:DEAD-box RNA Helicases / Q000235:genetics* / Q000378:metabolism; D003902:Detergents / Q000737:chemistry; D004722:Endoribonucleases / Q000235:genetics* / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D005947:Glucose / Q000172:deficiency; D009997:Osmotic Pressure; D011132:Polyribosomes / Q000235:genetics / Q000378:metabolism; D066329:Protein Aggregates; D014176:Protein Biosynthesis; D039381:RNA Cap-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D020871:RNA Stability; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism / Q000648:ultrastructure; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000378:metabolism; D013312:Stress, Physiological / Q000235:genetics; D013395:Sucrose
chemical_list: D003902:Detergents; D066329:Protein Aggregates; D039381:RNA Cap-Binding Proteins; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins; D013395:Sucrose; C100868:DCP1 protein, S cerevisiae; C400572:DCP2 protein, S cerevisiae; D004722:Endoribonucleases; C445083:DHH1 protein, S cerevisiae; D053487:DEAD-box RNA Helicases; D005947:Glucose