RSR1, a ras-like gene homologous to Krev-1 (smg21A/rap1A): role in the development of cell polarity and interactions with the Ras pathway in Saccharomyces cerevisiae

The Saccharomyces cerevisiae ras-like gene RSR1 is particularly closely related to the mammalian gene Krev-1 (also known as smg21A and rap1A). RSR1 was originally isolated as a multicopy suppressor of a cdc24 mutation, which causes an inability to bud or establish cell polarity. Deletion of RSR1 itself does not affect growth but causes a randomization of bud position. We have now constructed mutant alleles of RSR1 encoding proteins with substitutions of Val for Gly at position 12 (analogous to constitutively activated Ras proteins) or Asn for Lys at position 16 (analogous to a dominant-negative Ras protein). rsr1Val-12 could not restore a normal budding pattern to an rsr1 deletion strain but could suppress a cdc24 mutation when overexpressed. rsr1Asn-16 could randomize the budding pattern of a wild-type strain even in low copy number but was not lethal even in high copy number. These and other results suggest that Rsr1p functions only in bud site selection and not in subsequent events of polarity establishment and bud formation, that this function involves a cycling between GTP-bound and GDP-bound forms of the protein, and that the suppression of cdc24 involves direct interaction between Rsr1p[GTP] and Cdc24p. Functional homology between Rsr1p and Krev-1 p21 was suggested by the observations that expression of the latter protein in yeast cells could both suppress a cdc24 mutation and randomize the budding pattern of wild-type cells. As Krev-1 overexpression can suppress ras-induced transformation of mammalian cells, we looked for effects of RSR1 on the S. cerevisiae Ras pathway. Although no suppression of the activated RAS2Val-19 allele was observed, overexpression of rsr1Val-12 suppressed the lethality of strains lacking RAS gene function, apparently through a direct activation of adenyl cyclase. This interaction of Rsr1p with the effector of Ras in S. cerevisiae suggests that Krev-1 may revert ras-induced transformation of mammalian cells by affecting the interaction of ras p21 with its effector.

[1]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[2]  I. Herskowitz,et al.  Genetic control of bud site selection in yeast by a set of gene products that constitute a morphogenetic pathway , 1991, Cell.

[3]  J. Broach,et al.  The CDC25 protein of Saccharomyces cerevisiae promotes exchange of guanine nucleotides bound to ras , 1991, Molecular and cellular biology.

[4]  F. Tamanoi,et al.  IRA2, an upstream negative regulator of RAS in yeast, is a RAS GTPase-activating protein. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[5]  Frank McCormick,et al.  The GTPase superfamily: a conserved switch for diverse cell functions , 1990, Nature.

[6]  M. Wigler,et al.  Biological activity of the mammalian RAP genes in yeast. , 1990, Cell regulation.

[7]  M. Wigler,et al.  GAPs in understanding Ras , 1990, Nature.

[8]  A. Hall,et al.  The cellular functions of small GTP-binding proteins. , 1990, Science.

[9]  K. Matsumoto,et al.  IRA2, a second gene of Saccharomyces cerevisiae that encodes a protein with a domain homologous to mammalian ras GTPase-activating protein. , 1990, Molecular and cellular biology.

[10]  A. Wittinghofer,et al.  Inhibition of GTPase activating protein stimulation of Ras-p21 GTPase by the Krev-1 gene product. , 1990, Science.

[11]  J. Pringle,et al.  CDC42 and CDC43, two additional genes involved in budding and the establishment of cell polarity in the yeast Saccharomyces cerevisiae , 1990, The Journal of cell biology.

[12]  J. Pringle,et al.  Molecular characterization of CDC42, a Saccharomyces cerevisiae gene involved in the development of cell polarity , 1990, The Journal of cell biology.

[13]  Alan Hall,et al.  ras and GAP—Who's controlling whom? , 1990, Cell.

[14]  H. Kitayama,et al.  Genetic analysis of the Kirsten-ras-revertant 1 gene: potentiation of its tumor suppressor activity by specific point mutations. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[15]  M. Jacquet,et al.  Enhancement of the GDP-GTP exchange of RAS proteins by the carboxyl-terminal domain of SCD25. , 1990, Science.

[16]  A. Levitzki.  GTP-GDP exchange proteins. , 1990, Science.

[17]  T. Sasaki,et al.  Inhibition of the ras p21 GTPase-activating protein-stimulated GTPase activity of c-Ha-ras p21 by smg p21 having the same putative effector domain as ras p21s. , 1990, The Journal of biological chemistry.

[18]  M. Nakafuku,et al.  S. cerevisiae genes IRA1 and IRA2 encode proteins that may be functionally equivalent to mammalian ras GTPase activating protein , 1990, Cell.

[19]  J. Pringle,et al.  Multicopy suppression of the cdc24 budding defect in yeast by CDC42 and three newly identified genes including the ras-related gene RSR1. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[20]  M. Wigler,et al.  Genetic analysis of mammalian GAP expressed in yeast , 1989, Cell.

[21]  M. Marshall,et al.  The ras oncogene--an important regulatory element in lower eucaryotic organisms , 1989, Microbiological reviews.

[22]  G. Sprague,,et al.  Pheromones and pheromone receptors are the primary determinants of mating specificity in the yeast Saccharomyces cerevisiae. , 1989, Genetics.

[23]  M. Wigler,et al.  Dominant yeast and mammalian RAS mutants that interfere with the CDC25-dependent activation of wild-type RAS in Saccharomyces cerevisiae , 1989, Molecular and cellular biology.

[24]  H. Kitayama,et al.  A ras-related gene with transformation suppressor activity , 1989, Cell.

[25]  Frank McCormick,et al.  ras GTPase activating protein: Signal transmitter and signal terminator , 1989, Cell.

[26]  Y. Takai,et al.  A novel small molecular weight GTP-binding protein with the same putative effector domain as the ras proteins in bovine brain membranes. Purification, determination of primary structure, and characterization. , 1988, The Journal of biological chemistry.

[27]  B. Olofsson,et al.  Human cDNAs rap1 and rap2 homologous to the Drosophila gene Dras3 encode proteins closely related to ras in the 'effector' region. , 1988, Oncogene.

[28]  H. Bourne.  Do GTPases direct membrane traffic in secretion? , 1988, Cell.

[29]  D. Lowy,et al.  Guanosine triphosphatase activating protein (GAP) interacts with the p21 ras effector binding domain. , 1988, Science.

[30]  C. Marshall,et al.  The cytoplasmic protein GAP is implicated as the target for regulation by the ras gene product , 1988, Nature.

[31]  K. Arai,et al.  Isolation of a second yeast Saccharomyces cerevisiae gene (GPA2) coding for guanine nucleotide-binding regulatory protein: studies on its structure and possible functions. , 1988, Proceedings of the National Academy of Sciences of the United States of America.

[32]  K. Matsumoto,et al.  Dual regulation of the expression of the polyubiquitin gene by cyclic AMP and heat shock in yeast. , 1988, The EMBO journal.

[33]  E. Scolnick,et al.  Identification of guanine nucleotides bound to ras-encoded proteins in growing yeast cells. , 1987, The Journal of biological chemistry.

[34]  K. Tatchell,et al.  Characterization of Saccharomyces cerevisiae genes encoding subunits of cyclic AMP-dependent protein kinase , 1987, Molecular and cellular biology.

[35]  Michael Wigler,et al.  Three different genes in S. cerevisiae encode the catalytic subunits of the cAMP-dependent protein kinase , 1987, Cell.

[36]  E. Scolnick,et al.  Regulatory function of the Saccharomyces cerevisiae RAS C-terminus , 1987, Molecular and cellular biology.

[37]  M. Wigler,et al.  The S. cerevisiae CDC25 gene product regulates the RAS/adenylate cyclase pathway , 1987, Cell.

[38]  L. C. Robinson,et al.  CDC25: a component of the RAS-adenylate cyclase pathway in Saccharomyces cerevisiae. , 1987, Science.

[39]  R. Axel,et al.  Characterization of two members of the rho gene family from the yeast Saccharomyces cerevisiae. , 1987, Proceedings of the National Academy of Sciences of the United States of America.

[40]  E. Scolnick,et al.  Identification of effector residues and a neutralizing epitope of Ha-ras-encoded p21. , 1986, Proceedings of the National Academy of Sciences of the United States of America.

[41]  E. Scolnick,et al.  Mutant ras-encoded proteins with altered nucleotide binding exert dominant biological effects. , 1986, Proceedings of the National Academy of Sciences of the United States of America.

[42]  M. Holland,et al.  Isolation and characterization of yeast strains carrying mutations in the glyceraldehyde-3-phosphate dehydrogenase genes. , 1985, The Journal of biological chemistry.

[43]  M. Wigler,et al.  DNA sequence and characterization of the S. cerevisiae gene encoding adenylate cyclase , 1985, Cell.

[44]  K. Arai,et al.  Nucleotide sequences of STE2 and STE3, cell type‐specific sterile genes from Saccharomyces cerevisiae , 1985, The EMBO journal.

[45]  L. C. Robinson,et al.  Mammalian and yeast ras gene products: biological function in their heterologous systems. , 1985, Science.

[46]  M. Wigler,et al.  In yeast, RAS proteins are controlling elements of adenylate cyclase , 1985, Cell.

[47]  M. Wigler,et al.  Genetic analysis of yeast RAS1 and RAS2 genes , 1984, Cell.

[48]  M. Wigler,et al.  Genes in S. cerevisiae encoding proteins with domains homologous to the mammalian ras proteins , 1984, Cell.

[49]  K. Murata,et al.  Transformation of intact yeast cells treated with alkali cations. , 1984, Journal of bacteriology.

[50]  J. Pringle,et al.  Roles of the CDC24 gene product in cellular morphogenesis during the Saccharomyces cerevisiae cell cycle , 1981, The Journal of cell biology.

[51]  S. H. Lillie,et al.  Reserve carbohydrate metabolism in Saccharomyces cerevisiae: responses to nutrient limitation , 1980, Journal of bacteriology.

[52]  D. Botstein,et al.  Sterile host yeasts (SHY): a eukaryotic system of biological containment for recombinant DNA experiments. , 1979, Gene.

[53]  J. Broach,et al.  Transformation in yeast: development of a hybrid cloning vector and isolation of the CAN1 gene. , 1979, Gene.

[54]  M. Holland,et al.  The primary structure of a glyceraldehyde-3-phosphate dehydrogenase gene from Saccharomyces cerevisiae. , 1979, The Journal of biological chemistry.

[55]  F. Sanger,et al.  DNA sequencing with chain-terminating inhibitors. , 1977, Proceedings of the National Academy of Sciences of the United States of America.

[56]  L. Hartwell,et al.  Genetic control of the cell division cycle in yeast. , 1974, Science.

[57]  L. Hartwell,et al.  Genetic Control of the Cell Division Cycle in Yeast: V. Genetic Analysis of cdc Mutants. , 1973, Genetics.

[58]  B. Haarer,et al.  Fluorescence microscopy methods for yeast. , 1989, Methods in cell biology.

[59]  D. Botstein,et al.  A Saccharomyces cerevisiae genomic plasmid bank based on a centromere-containing shuttle vector. , 1987, Gene.

[60]  M. Barbacid.  ras genes. , 1987, Annual review of biochemistry.

[61]  C. Yanisch-Perron,et al.  Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. , 1985, Gene.

[62]  Gerald R. Fink,et al.  Methods in Yeast Genetics: Laboratory Manual , 1981 .

[63]  Kunihiro Matsumoto,et al.  IRA ] , an Inhibitory Regulator of the RAS-Cyclic AMP Pathway in Saccharomyces cerevisiae , 2022 .