Tor proteins and protein phosphatase 2A reciprocally regulate Tap42 in controlling cell growth in yeast

Tor proteins, homologous to DNA‐dependent protein kinases, participate in a signal transduction pathway in yeast that regulates protein synthesis and cell wall expansion in response to nutrient availability. The anti‐inflammatory drug rapamycin inhibits yeast cell growth by inhibiting Tor protein signaling. This leads to diminished association of a protein, Tap42, with two different protein phosphatase catalytic subunits; one encoded redundantly by PPH21 and PPH22, and one encoded by SIT4. We show that inactivation of either Cdc55 or Tpd3, which regulate Pph21/22 activity, results in rapamycin resistance and that this resistance correlates with an increased association of Tap42 with Pph21/22. Furthermore, we show Tor‐dependent phosphorylation of Tap42 both in vivo and in vitro and that this phosphorylation is rapamycin sensitive. Inactivation of Cdc55 or Tpd3 enhances in vivo phosphorylation of Tap42. We conclude that Tor phosphorylates Tap42 and that phosphorylated Tap42 effectively competes with Cdc55/Tpd3 for binding to the phosphatase 2A catalytic subunit. Furthermore, Cdc55 and Tpd3 promote dephosphorylation of Tap42. Thus, Tor stimulates growth‐promoting association of Tap42 with Pph21/22 and Sit4, while Cdc55 and Tpd3 inhibit this association both by direct competition and by dephosphorylation of Tap42. These results establish Tap42 as a target of Tor and add further refinement to the Tor signaling pathway.

[1]  A. Schmidt,et al.  The TOR nutrient signalling pathway phosphorylates NPR1 and inhibits turnover of the tryptophan permease , 1998, The EMBO journal.

[2]  A. Shaw,et al.  The 14-3-3 proteins positively regulate rapamycin-sensitive signaling , 1998, Current Biology.

[3]  Y. Ohya,et al.  The Rho1 effector Pkc1, but not Bni1, mediates signalling from Tor2 to the actin cytoskeleton , 1998, Current Biology.

[4]  P. Graves,et al.  14-3-3 Proteins Are Required for Maintenance of Raf-1 Phosphorylation and Kinase Activity , 1998, Molecular and Cellular Biology.

[5]  S. Schreiber,et al.  Alpha 4 associates with protein phosphatases 2A, 4, and 6. , 1998, Biochemical and biophysical research communications.

[6]  R. Roth,et al.  Evidence of insulin-stimulated phosphorylation and activation of the mammalian target of rapamycin mediated by a protein kinase B signaling pathway. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[7]  J. Avruch,et al.  Amino Acid Sufficiency and mTOR Regulate p70 S6 Kinase and eIF-4E BP1 through a Common Effector Mechanism* , 1998, The Journal of Biological Chemistry.

[8]  C. Berset,et al.  The TOR (target of rapamycin) signal transduction pathway regulates the stability of translation initiation factor eIF4G in the yeast Saccharomyces cerevisiae. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[9]  S. Snyder,et al.  RAFT1 phosphorylation of the translational regulators p70 S6 kinase and 4E-BP1. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[10]  S. Snyder,et al.  RAFT 1 phosphorylation of the translational regulators p 70 S 6 kinase and 4 EBP 1 , 1998 .

[11]  T. Haystead,et al.  The Mammalian Target of Rapamycin Phosphorylates Sites Having a (Ser/Thr)-Pro Motif and Is Activated by Antibodies to a Region near Its COOH Terminus , 1997, The Journal of Biological Chemistry.

[12]  G. Thomas,et al.  TOR signalling and control of cell growth. , 1997, Current opinion in cell biology.

[13]  Jun Wu,et al.  B cell receptor-associated protein α4 displays rapamycin-sensitive binding directly to the catalytic subunit of protein phosphatase 2A , 1997 .

[14]  Christine C. Hudson,et al.  Phosphorylation of the translational repressor PHAS-I by the mammalian target of rapamycin. , 1997, Science.

[15]  R. Pearson,et al.  Rapamycin suppresses 5′TOP mRNA translation through inhibition of p70s6k , 1997, The EMBO journal.

[16]  Y. Shu,et al.  Molecular genetic analysis of Rts1p, a B' regulatory subunit of Saccharomyces cerevisiae protein phosphatase 2A , 1997, Molecular and cellular biology.

[17]  Marc Bickle,et al.  The Yeast Phosphatidylinositol Kinase Homolog TOR2 Activates RHO1 and RHO2 via the Exchange Factor ROM2 , 1997, Cell.

[18]  M. Hall,et al.  TOR2 is required for organization of the actin cytoskeleton in yeast. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[19]  K. Arndt,et al.  Nutrients, via the Tor proteins, stimulate the association of Tap42 with type 2A phosphatases. , 1996, Genes & development.

[20]  K. Arndt,et al.  The SAP, a new family of proteins, associate and function positively with the SIT4 phosphatase , 1996, Molecular and cellular biology.

[21]  C. Proud.  p70 S6 kinase: an enigma with variations. , 1996, Trends in biochemical sciences.

[22]  A. Gingras,et al.  4E-BP1 phosphorylation is mediated by the FRAP-p70s6k pathway and is independent of mitogen-activated protein kinase. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[23]  P. Allen,et al.  Interaction of 14-3-3 with Signaling Proteins Is Mediated by the Recognition of Phosphoserine , 1996, Cell.

[24]  A. Gingras,et al.  Rapamycin blocks the phosphorylation of 4E‐BP1 and inhibits cap‐dependent initiation of translation. , 1996, The EMBO journal.

[25]  M. Hall,et al.  The TOR signalling pathway and growth control in yeast. , 1996, Biochemical Society transactions.

[26]  I. Stansfield,et al.  An MBoC Favorite: TOR controls translation initiation and early G1 progression in yeast , 2012, Molecular biology of the cell.

[27]  J. Heitman,et al.  FKBP12‐rapamycin target TOR2 is a vacuolar protein with an associated phosphatidylinositol‐4 kinase activity. , 1995, The EMBO journal.

[28]  J. Heitman,et al.  TOR Mutations Confer Rapamycin Resistance by Preventing Interaction with FKBP12-Rapamycin (*) , 1995, The Journal of Biological Chemistry.

[29]  S. Schreiber,et al.  Control of p70 S6 kinase by kinase activity of FRAP in vivo , 1995, Nature.

[30]  Stuart L. Schreiber,et al.  TOR kinase domains are required for two distinct functions, only one of which is inhibited by rapamycin , 1995, Cell.

[31]  S. Schreiber,et al.  Erratum: Control of p70 S6 kinase by kinase activity of FRAP in vivo (Nature (1995) 377 (441-446)) , 1995 .

[32]  M. Mclaughlin,et al.  Interaction between FKBP12-rapamycin and TOR involves a conserved serine residue. , 1994, The Journal of biological chemistry.

[33]  J. Kunz,et al.  TOR1 and TOR2 are structurally and functionally similar but not identical phosphatidylinositol kinase homologues in yeast. , 1994, Molecular biology of the cell.

[34]  J. Kunz,et al.  Target of rapamycin in yeast, TOR2, is an essential phosphatidylinositol kinase homolog required for G1 progression , 1993, Cell.

[35]  K. Arndt,et al.  SIT4 protein phosphatase is required for the normal accumulation of SWI4, CLN1, CLN2, and HCS26 RNAs during late G1. , 1992, Genes & development.

[36]  J. Broach,et al.  Inactivation of the protein phosphatase 2A regulatory subunit A results in morphological and transcriptional defects in Saccharomyces cerevisiae , 1992, Molecular and cellular biology.

[37]  V. Simanis.  Guide to yeast genetics and molecular biology: edited by Christine Guthrie and Gerald R. Fink, Academic Press, 1991. £35.50/$49.95 (xxi + 933 pages) ISBN 0 12 310670 2 , 1992 .

[38]  M. Goebl,et al.  CDC55, a Saccharomyces cerevisiae gene involved in cellular morphogenesis: identification, characterization, and homology to the B subunit of mammalian type 2A protein phosphatase , 1991, Molecular and cellular biology.

[39]  H. Ronne,et al.  Protein phosphatase 2A in Saccharomyces cerevisiae: effects on cell growth and bud morphogenesis , 1991, Molecular and cellular biology.

[40]  J. Heitman,et al.  Targets for cell cycle arrest by the immunosuppressant rapamycin in yeast , 1991, Science.

[41]  K. Arndt,et al.  The SIT4 protein phosphatase functions in late G1 for progression into S phase , 1991, Molecular and cellular biology.

[42]  J. Heitman,et al.  FK 506-binding protein proline rotamase is a target for the immunosuppressive agent FK 506 in Saccharomyces cerevisiae. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[43]  Janina Maier,et al.  Guide to yeast genetics and molecular biology. , 1991, Methods in enzymology.

[44]  P. Cohen,et al.  Saccharomyces cerevisiae protein phosphatase 2A performs an essential cellular function and is encoded by two genes. , 1990, The EMBO journal.

[45]  G M Rubin,et al.  The nucleotide sequence of Saccharomyces cerevisiae 5.8 S ribosomal ribonucleic acid. , 1973, The Journal of biological chemistry.

[46]  M. Rogers,et al.  Amphibian ribosomal ribonucleic acids. , 1972, The Biochemical journal.