title: Protein Kinase C Controls Binding of Igo/ENSA Proteins to Protein Phosphatase 2A in Budding Yeast.
abstract: Protein phosphatase 2A (PP2A) plays important roles in controlling mitosis in all eukaryotic cells. The form of PP2A that controls mitosis is associated with a conserved regulatory subunit that is called B55 in vertebrates and Cdc55 in budding yeast. The activity of this form of PP2A can be inhibited by binding of conserved Igo/ENSA proteins. Although the mechanisms that activate Igo/ENSA to bind and inhibit PP2A are well understood, little is known about how Igo/Ensa are inactivated. Here, we have analyzed regulation of Igo/ENSA in the context of a checkpoint pathway that links mitotic entry to membrane growth in budding yeast. Protein kinase C (Pkc1) relays signals in the pathway by activating PP2ACdc55We discovered that constitutively active Pkc1 can drive cells through a mitotic checkpoint arrest, which suggests that Pkc1-dependent activation of PP2ACdc55plays a critical role in checkpoint signaling. We therefore used mass spectrometry to determine how Pkc1 modifies the PP2ACdc55complex. This revealed that Pkc1 induces changes in the phosphorylation of multiple subunits of the complex, as well as dissociation of Igo/ENSA. Pkc1 directly phosphorylates Cdc55 and Igo/ENSA, and phosphorylation site mapping and mutagenesis indicate that phosphorylation of Cdc55 contributes to Igo/ENSA dissociation. Association of Igo2 with PP2ACdc55is regulated during the cell cycle, yet mutation of Pkc1-dependent phosphorylation sites on Cdc55 and Igo2 did not cause defects in mitotic progression. Together, the data suggest that Pkc1 controls PP2ACdc55by multiple overlapping mechanisms.
authors.affiliation: From the Department of Molecular, Cell and Developmental Biology, University of California at Santa Cruz, Santa Cruz, California 95064.
authors.forename: Vu
authors.initials: V
authors.lastname: Thai
authors.affiliation: the Department of Biochemistry, Weill Cornell Medical College, New York, New York 10021, and.
authors.forename: Noah
authors.initials: N
authors.lastname: Dephoure
authors.affiliation: From the Department of Molecular, Cell and Developmental Biology, University of California at Santa Cruz, Santa Cruz, California 95064.
authors.forename: Amit
authors.initials: A
authors.lastname: Weiss
authors.affiliation: From the Department of Molecular, Cell and Developmental Biology, University of California at Santa Cruz, Santa Cruz, California 95064.
authors.forename: Jacqueline
authors.initials: J
authors.lastname: Ferguson
authors.affiliation: From the Department of Molecular, Cell and Developmental Biology, University of California at Santa Cruz, Santa Cruz, California 95064.
authors.forename: Ricardo
authors.initials: R
authors.lastname: Leitao
authors.affiliation: the Department of Cell Biology, Harvard Medical School, Boston, Massachusetts 02115.
authors.forename: Steven P
authors.initials: SP
authors.lastname: Gygi
authors.affiliation: From the Department of Molecular, Cell and Developmental Biology, University of California at Santa Cruz, Santa Cruz, California 95064, dkellogg@ucsc.edu.
authors.forename: Douglas R
authors.identifier: 0000-0002-5050-2194
authors.initials: DR
authors.lastname: Kellogg
mesh_terms: D000595:Amino Acid Sequence; D018797:Cell Cycle Proteins / Q000032:analysis / Q000378:metabolism*; D008958:Models, Molecular; D010766:Phosphorylation; D011485:Protein Binding; D011493:Protein Kinase C / Q000032:analysis / Q000378:metabolism*; D054648:Protein Phosphatase 2 / Q000032:analysis / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000166:cytology* / Q000254:growth & development / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000032:analysis / Q000378:metabolism*; D016415:Sequence Alignment
chemical_list: C071246:CDC55 protein, S cerevisiae; D018797:Cell Cycle Proteins; C549818:Igo1 protein, S cerevisiae; C549819:Igo2 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C490702:PKC1 protein, S cerevisiae; D011493:Protein Kinase C; D054648:Protein Phosphatase 2
keywords: B55; Cdc55; ENSA; Igo2; Pkc1; Saccharomyces cerevisiae; cell cycle; cell growth; protein kinase C (PKC); protein phosphatase 2 (PP2A)