The Byr2 Kinase Translocates to the Plasma Membrane in a Ras1-Dependent Manner

Bauman, Patricia; Cheng, Qiu-chen; Albright, Charles F.

doi:10.1006/bbrc.1998.8292

Cited by 22 publications

(18 citation statements)

References 51 publications

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“…S2A in the supplemental material) that resemble vacuoles. The Ras1-Byr2 mating pathway is tightly regulated spatially: Byr2 has been shown to translocate to the PM at the onset of sexual differentiation (32). Strikingly, when GFP-Ras1 was examined during sexual differentiation, it was essentially undetectable at the PM in all of the isolated mutants (Fig.…”

Section: Resultsmentioning

confidence: 92%

Regulation of Ras Localization and Cell Transformation by Evolutionarily Conserved Palmitoyltransferases

Young

Zheng

Wilkins

et al. 2014

Molecular and Cellular Biology

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e Ras can act on the plasma membrane (PM) to mediate extracellular signaling and tumorigenesis. To identify key components controlling Ras PM localization, we performed an unbiased screen to seek Schizosaccharomyces pombe mutants with reduced PM Ras. Five mutants were found with mutations affecting the same gene, S. pombe erf2 (sp-erf2), encoding sp-Erf2, a palmitoyltransferase, with various activities. sp-Erf2 localizes to the trans-Golgi compartment, a process which is mediated by its third transmembrane domain and the Erf4 cofactor. In fission yeast, the human ortholog zDHHC9 rescues the phenotypes of sp-erf2 null cells. In contrast, expressing zDHHC14, another sp-Erf2-like human protein, did not rescue Ras1 mislocalization in these cells. Importantly, ZDHHC9 is widely overexpressed in cancers. Overexpressing ZDHHC9 promotes, while repressing it diminishes, Ras PM localization and transformation of mammalian cells. These data strongly demonstrate that sp-Erf2/zDHHC9 palmitoylates Ras proteins in a highly selective manner in the trans-Golgi compartment to facilitate PM targeting via the transGolgi network, a role that is most certainly critical for Ras-driven tumorigenesis. Mammals have three RAS genes, HRAS, NRAS, and KRAS, which encode membrane-bound GTPases that control a wide range of signal transduction pathways critical for growth, differentiation, and cytoskeleton remodeling (1). The best-known Ras function is to mediate growth factor signaling that occurs at the plasma membrane (PM), and deregulation of this and other Ras functions can promote tumor formation (2, 3). While all Ras proteins are farnesylated to gain general membrane affinity, a second signal is needed to specifically localize to the PM. For HRas and NRas, this signal is provided by palmitoylation at cysteine residues immediately upstream of the C-terminal CAAX motif. This reaction can be catalyzed by protein acyltransferases (PATs) that contain a DHHC domain rich with aspartate, histidine, and cysteine residues. These DHHC-PATs are expected to localize in the Golgi complex to allow the delivery of palmitoylated Ras proteins to the PM through the trans-Golgi system (4) although this concept has not been thoroughly demonstrated in vivo.Humans have 24 DHHC-PATs, but it remains unresolved which of these control palmitoylation of Ras proteins in vivo. In contrast, the Ras pathways are considerably simpler in yeasts. Early studies from the budding yeast Saccharomyces cerevisiae, which has seven DHHC-PATs and two Ras proteins, suggest that only S. cerevisiae Erf2 (sc-Erf2) controls palmitoylation of its Ras proteins (5, 6). By sequence alignment to find putative orthologs, zDHHC9 was predicted to be a human Ras PAT (7). Although zDHHC9 can catalyze Ras palmitoylation in vitro, zDHHC9 cannot rescue sc-erf2 mutant phenotypes in budding yeast (8), and until now there has been no evidence that zDHHC9 or other human DHHC-PATs can control Ras localization in mammalian cells. Furthermore, while ectopically expressed zDHHC9 localizes to the Golgi compl...

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Section: Resultsmentioning

confidence: 92%

Regulation of Ras Localization and Cell Transformation by Evolutionarily Conserved Palmitoyltransferases

Young

Zheng

Wilkins

et al. 2014

Molecular and Cellular Biology

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“…Earlier work has shown that Byr2 recognizes membraneassociated Ras-1 in its active GTP bound state via a Ras- binding domain (28,40). Through Ras-1 association and by interactions with other proteins such as Shk1, Byr2 may become converted to an "open conformation," thereby relieving the autoinhibition of its kinase activity (30).…”

Section: Fig 3 Equilibrium Sedimentation Of Sam Domainsmentioning

confidence: 99%

Oligomerization-dependent Association of the SAM Domains from Schizosaccharomyces pombe Byr2 and Ste4

Ramachander

Kim

Phillips

et al. 2002

Journal of Biological Chemistry

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SAM domains (also known as Pointed, SPM, and HLH domains) are frequently found in eukaryotic regulatory proteins ranging from receptor tyrosine kinases to transcription factors (1-3). Structures of several SAM domains reveal a common tertiary fold but show a diverse array of oligomeric states and binding schemes (4 -11). Some SAM domains, such as that from the Ets family transcription factor TEL, can self-associate to form an open-ended polymeric structure (10), whereas the closely related Ets-1, GABP␣, and Erg SAM domains are monomeric (4, 12, 13). The SAM domains from Eph receptor tyrosine kinases can either be monomeric or dimeric or may possibly form an extended oligomeric structure (7,9,14). SAM domains have also been described in interactions with non-SAM domain-containing proteins. For example, the SAM domain of BAR (46), a protein involved in the regulation of apoptosis, associates with both Bcl-2 and Bcl-XL (13). Cdk10, a member of the Cdc2 family of kinases, binds the SAM domain of Ets-2 and thereby regulates the activity of this transcription factor (15). The mitogen-activated protein kinase Erk2 docks on the SAM domain of Ets-1, enhancing the kinetics of phosphorylation at an adjacent N-terminal target site within this transcription factor (16). Although several complexes between nonidentical SAM domains have been described like TEL/TEL2 (17-23), and Yan/Mae (24) Scm/ph (3,11,25), their recognition mechanisms have not yet been characterized. Here we investigate one example of a hetero-SAM domain interaction that occurs between the Byr2 and Ste4 proteins in the fission yeast Schizosaccharomyces pombe.Sexual differentiation in S. pombe is controlled via a mitogen-activated protein kinase pathway that includes Ste4 and Byr2 (26). Byr2 is a mitogen-activated protein kinase kinase kinase that is activated by interactions with both . The SAM domain of Byr2 has previously been shown to bind to the N-terminal 160 amino acids of Ste4, a region containing a SAM domain followed immediately by a putative leucine zipper (Ste4-LZ) domain. A speculative model of Byr2 activation has therefore emerged in which Byr2 and Ste4 interact via their SAM domains, leading to oligomerization of Byr2 by virtue of the Ste4 leucine zipper domain (30). Here we find that although the two SAM domains do bind to each other, the role of the Ste4-LZ domain is not to oligomerize Byr2. Instead, the leucine zipper domain of Ste4 trimerizes, thereby displaying three SAM domains that together bind a single Byr2-SAM domain with high affinity. EXPERIMENTAL PROCEDURESSte4 and Byr2 Constructs-The region of the Byr2 gene encompassing its SAM domain (amino acids 1-70; SPBC1D7.05 in the S. pombe GeneDB, www.genedb.org/genedb/pombe/index.jsp) was PCR-amplified from a S. pombe cDNA library and cloned into a modified pET-3c (Novagen) expression vector containing a C-terminal six-histidine tag. The expressed protein sequence comprised amino acids 1-70 of Byr2, followed by RDHHHHHH. The DNA sequences encoding the SAM domain (amino acids 9 -72) and the...

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“…Moreover, the farthest-upstream component, Kpp4/Ubc4, contains a RA domain that is conserved in proteins interacting with Ras proteins (3; Müller et al, submitted). In S. pombe, Ras1 regulates the Ubc4 homologue, Byr2, by interacting with the RA domain of Byr2 MAPKK kinase, and this interaction leads to the translocation of Byr2 to the plasma membrane (8,45,63). Recently, the ras2 gene of U. maydis was found to act upstream of kpp2/ubc3 (42).…”

Section: Discussionmentioning

confidence: 99%

Guanyl Nucleotide Exchange Factor Sql2 and Ras2 Regulate Filamentous Growth in Ustilago maydis

et al. 2003

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The cyclic AMP (cAMP)-signaling pathway regulates cell morphology and plays a crucial role during pathogenic development of the plant-pathogenic fungus Ustilago maydis. Strains lacking components of this signaling pathway, such as the G␣-subunit Gpa3 or the adenylyl cyclase Uac1, are nonpathogenic and grow filamentously. On the other hand, strains exhibiting an activated cAMP pathway due to a dominant-active allele of gpa3 display a glossy colony phenotype and are unable to proliferate in plant tumors. Here we present the identification of sql2 as a suppressor of the glossy colony phenotype of a gpa3 Q206L strain. sql2 encodes a protein with similarity to CDC25-like guanine nucleotide exchange factors, which are known to act on Ras proteins. Overexpression of sql2 leads to filamentous growth that cannot be suppressed by exogenous cAMP, suggesting that Sql2 does not act upstream of Uac1. To gain more insight in signaling processes regulated by Sql2, we isolated two genes encoding Ras proteins. Expression of dominant active alleles of ras1 and ras2 showed that Ras2 induces filamentous growth while Ras1 does not affect cell morphology but elevates pheromone gene expression. These results indicate that Ras1 and Ras2 fulfill different functions in U. maydis. Moreover, observed similarities between the filaments induced by sql2 and ras2 suggest that Sql2 is an activator of Ras2. Interestingly, sql2 deletion mutants are affected in pathogenic development but not in mating, indicating a specific function of sql2 during pathogenesis.

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The Byr2 Kinase Translocates to the Plasma Membrane in a Ras1-Dependent Manner

Cited by 22 publications

References 51 publications

Regulation of Ras Localization and Cell Transformation by Evolutionarily Conserved Palmitoyltransferases

Regulation of Ras Localization and Cell Transformation by Evolutionarily Conserved Palmitoyltransferases

Oligomerization-dependent Association of the SAM Domains from Schizosaccharomyces pombe Byr2 and Ste4

Guanyl Nucleotide Exchange Factor Sql2 and Ras2 Regulate Filamentous Growth in Ustilago maydis

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