title: Genetic and biochemical interactions between SCP160 and EAP1 in yeast.
abstract: Scp160p is a multiple KH-domain RNA-binding protein in yeast known to associate with polyribosomes as an mRNP component, although its biological role remains unclear. As a genetic approach to examine Scp160p function, we applied an ethyl methanesulfonate (EMS) screen for loci synthetically lethal with scp160 loss, and identified a single candidate gene, EAP1, whose protein product functions in translation as an eIF4E-binding protein, with additional uncharacterized spindle pole body functions. To reconfirm scp160/eap1 synthetic lethality, we constructed a strain null for both genes, supported by an SCP160 maintenance plasmid. We used this strain to establish a quantitative assay for both Scp160p and Eap1p functions in vivo, and applied this assay to demonstrate that Y109A EAP1, a previously described allele of EAP1 that cannot bind eIF4E, is markedly impaired with regard to its SCP160-related activity. In addition, we explored the possibility of physical interaction between Eap1p and Scp160p, and discovered that Eap1p associates with Scp160p-containing complexes in an RNA-dependent manner. Finally, we probed the impact of EAP1 loss on Scp160p, and vice versa, and found that loss of each gene resulted in a significant change in either the complex associations or subcellular distribution of the other protein. These results clearly support the hypothesis that Scp160p plays a role in translation, demonstrate that the interaction of SCP160 and EAP1 is biologically significant, and provide important tools for future studies of the in vivo functions of both genes.
authors.affiliation: Department of Biology, Emory College, Emory University School of Medicine, Room 325.2 Whitehead Building, 615 Michael Street, Atlanta, GA 30322, USA.
authors.forename: Bryce A
authors.initials: BA
authors.lastname: Mendelsohn
authors.forename: Ai-Min
authors.initials: AM
authors.lastname: Li
authors.forename: Claudia A
authors.initials: CA
authors.lastname: Vargas
authors.forename: Kristen
authors.initials: K
authors.lastname: Riehman
authors.forename: Alice
authors.initials: A
authors.lastname: Watson
authors.forename: Judith L
authors.initials: JL
authors.lastname: Fridovich-Keil
mesh_terms: D000483:Alleles; D002455:Cell Division / Q000187:drug effects / Q000235:genetics; D002462:Cell Membrane / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D005020:Ethyl Methanesulfonate / Q000494:pharmacology; D039561:Eukaryotic Initiation Factor-4E / Q000378:metabolism; D005804:Genes, Lethal / Q000235:genetics; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D010448:Peptide Initiation Factors / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D016601:RNA-Binding Proteins; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D039561:Eukaryotic Initiation Factor-4E; D008565:Membrane Proteins; D009687:Nuclear Proteins; D010448:Peptide Initiation Factors; D016601:RNA-Binding Proteins; C097262:SCP160 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C410795:eIF4E-associated protein Eap1; D005020:Ethyl Methanesulfonate