The Saccharomyces cerevisiae MET3 gene: Nucleotide sequence and relationship of the 5′ non-coding region to that of MET25

Cherest, Hélène; Kerjan, Pierre; Surdin-Kerjan, Yolande

doi:10.1007/bf00325699

Cited by 75 publications

(49 citation statements)

References 28 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The presence of this allele results in constitutively elevated levels of Ras/PKA signaling activity (30, 36). For most of the experiments in this study, we used an inducible version of this allele where RAS2 val19 was placed under the control of the promoter from the yeast MET3 gene (7,63,64). This promoter is repressed when cells are grown in medium containing methionine and induced when methionine is absent (7,54).…”

Section: Resultsmentioning

confidence: 99%

The Ras/cAMP-dependent Protein Kinase Signaling Pathway Regulates an Early Step of the Autophagy Process in Saccharomyces cerevisiae

Budovskaya

Stephan

Reggiori

et al. 2004

Journal of Biological Chemistry

194

168

View full text Add to dashboard Cite

When faced with nutrient deprivation, Saccharomyces cerevisiae cells enter into a nondividing resting state, known as stationary phase. The Ras/PKA (cAMPdependent protein kinase) signaling pathway plays an important role in regulating the entry into this resting state and the subsequent survival of stationary phase cells. The survival of these resting cells is also dependent upon autophagy, a membrane trafficking pathway that is induced upon nutrient deprivation. Autophagy is responsible for targeting bulk protein and other cytoplasmic constituents to the vacuolar compartment for their ultimate degradation. The data presented here demonstrate that the Ras/PKA signaling pathway inhibits an early step in autophagy because mutants with elevated levels of Ras/PKA activity fail to accumulate transport intermediates normally associated with this process. Quantitative assays indicate that these increased levels of Ras/PKA signaling activity result in an essentially complete block to autophagy. Interestingly, Ras/PKA activity also inhibited a related process, the cytoplasm to vacuole targeting (Cvt) pathway that is responsible for the delivery of a subset of vacuolar proteins in growing cells. These data therefore indicate that the Ras/PKA signaling pathway is not regulating a switch between the autophagy and Cvt modes of transport. Instead, it is more likely that this signaling pathway is controlling an activity that is required during the early stages of both of these membrane trafficking pathways. Finally, the data suggest that at least a portion of the Ras/PKA effects on stationary phase survival are the result of the regulation of autophagy activity by this signaling pathway.

show abstract

Section: Resultsmentioning

confidence: 99%

The Ras/cAMP-dependent Protein Kinase Signaling Pathway Regulates an Early Step of the Autophagy Process in Saccharomyces cerevisiae

Budovskaya

Stephan

Reggiori

et al. 2004

Journal of Biological Chemistry

194

168

View full text Add to dashboard Cite

show abstract

“…Interestingly, the HXGH motif is completely conserved in cloned bifunctional PAPS synthase proteins, i.e. the marine worm (5), mouse (4,17), human (3), guinea pig, 2 and fruit fly (6) as well as the ATP sulfurylases from fungi (18,19), plants (20,21), and yeast (22). Importantly, the ATP sulfurylase domain of bifunctional PAPS synthase is located in the COOH-terminal region of the protein (3), and it is in this segment of PAPS synthase that the HXGH motif is located.…”

Section: Discussionmentioning

confidence: 99%

Site-selected Mutagenesis of a Conserved Nucleotide Binding HXGH Motif Located in the ATP Sulfurylase Domain of Human Bifunctional 3′-Phosphoadenosine 5′-Phosphosulfate Synthase

Venkatachalam¹,

Fuda²,

Koonin³

et al. 1999

Journal of Biological Chemistry

View full text Add to dashboard Cite

3-Phosphoadenosine-5-phosphosulfate (PAPS) synthase is a bifunctional protein consisting of an NH 2 -terminal APS kinase and a COOH-terminal ATP sulfurylase. Both catalytic activities require ATP; the APS kinase domain involves cleavage of the ␤-␥ phosphodiester bond of ATP, whereas the ATP sulfurylase domain involves cleavage of the ␣-␤ phosphodiester bond of ATP. Previous mutational studies have suggested that ␤-␥ phosphodiesterase activity involves a highly conserved NTP-binding P-loop motif located in the adenosine-5-phosphosulfate kinase domain of PAPS synthases. Sequence alignment analysis of PAPS synthases and the superfamily of TagD-related nucleotidylyltransferases revealed the presence of a highly conserved HXGH motif in the ATP sulfurylase domain of PAPS synthases, a motif implicated in the ␣-␤ phosphodiesterase activity of cytidylyltransferases. Thus, site-selected mutagenesis of the HXGH motif in the ATP sulfurylase domain of human PAPS synthase (amino acids 425-428) was performed to examine this possibility. Either H425A or H428A mutation produced an inactive enzyme. In contrast, a N426K mutation resulted in increased enzymatic activity. A G427A single mutant resulted in only a modest 30% reduction in catalytic activity, whereas a G427A/H428A double mutant produced an inactive enzyme. These results suggest an important role for the HXGH histidines in the ATP sulfurylase activity of bifunctional PAPS synthase and support the hypothesis that the highly conserved HXGH motif found in the ATP sulfurylase domain of PAPS synthases is involved in ATP binding and ␣-␤ phosphodiesterase activity.In the course of sulfonation, inorganic sulfate must be activated prior to being transferred to an acceptor molecule (1), and the activated sulfate molecule is 3Ј-phosphoadenosine 5Ј-phosphosulfate (PAPS).1 The activation of inorganic sulfate to form PAPS results from the concerted action of two enzymes (1). The first step is catalyzed by ATP sulfurylase (ATP:sulfate adenylyltransferase, EC 2.7.7.4) and involves the reaction of inorganic sulfate with ATP to form adenosine 5Ј-phosphosulfate (APS) and inorganic pyrophosphate. This reaction results in the formation of a high energy phosphoric-sulfuric acid anhydride bond that is the chemical basis for sulfate activation (2). The second step involves APS reacting with another molecule of ATP to form PAPS and ADP and is catalyzed by APS kinase (ATP:adenylylsulfate 3Ј-phosphotransferase, EC 2.7.1.25). ATP sulfurylase and APS kinase cloned from bacteria, Fungi, yeast, and plants are found on separate polypeptide chains; however, in mammalian species, the marine worm, and the fruit fly, gene fusion has occurred, and the two enzymes are integral to bifunctional PAPS synthase. It has been clearly established that the NH 2 -terminal region of human (h) PAPS synthase constitutes the APS kinase domain, whereas the ATP sulfurylase domain is located in the COOH-terminal portion of this bifunctional protein (3). That is, the NH 2 -terminal 268 amino acids of hPAPS synthase (623 amino...

show abstract

“…Libraries of transmembrane domain (TMD) variants were made by PCR from SNC1 using degenerate PCR primers encoding random mixtures of phenylalanine, leucine, isoleucine, methionine, and valine. A plasmid expressing SNC1 from the methionine-repressible MET3 promoter (Cherest et al, 1987) was made by cloning the promoter (a gift from J. Holthuis) as an XhoI-EcoRI fragment in front of an SNC1 cDNA EcoRI-BamHI fragment in the vector pRS405. Alanine scanning mutants of SNC1 were made by PCR, which was facilitated by introducing silent ClaI, NarI and BglII sites at nucleotides 174, 229, and 264.…”

Section: Plasmidsmentioning

confidence: 99%

Specific Retrieval of the Exocytic SNARE Snc1p from Early Yeast Endosomes

Lewis

Nichols

Prescianotto-Baschong

et al. 2000

MBoC

330

515

View full text Add to dashboard Cite

Many endocytosed proteins in yeast travel to the vacuole, but some are recycled to the plasma membrane. We have investigated the recycling of chimeras containing green fluorescent protein (GFP) and the exocytic SNARE Snc1p. GFP-Snc1p moves from the cell surface to internal structures when Golgi function or exocytosis is blocked, suggesting continuous recycling via the Golgi. Internalization is mediated by a conserved cytoplasmic signal, whereas diversion from the vacuolar pathway requires sequences within and adjacent to the transmembrane domain. Delivery from the Golgi to the surface is also influenced by the transmembrane domain, but the requirements are much less specific. Recycling requires the syntaxins Tlg1p and Tlg2p but not Pep12p or proteins such as Vps4p and Vps5p that have been implicated in late endosome–Golgi traffic. Subtle changes to the recycling signal cause GFP-Snc1p to accumulate preferentially in punctate internal structures, although it continues to recycle to the surface. The internal GFP-Snc1p colocalizes with Tlg1p, and immunofluorescence and immunoelectron microscopy reveal structures that contain Tlg1p, Tlg2p, and Kex2p but lack Pep12p and Sec7p. We propose that these represent early endosomes in which sorting of Snc1p and late Golgi proteins occurs, and that transport can occur directly from them to the Golgi apparatus.

show abstract

The Saccharomyces cerevisiae MET3 gene: Nucleotide sequence and relationship of the 5′ non-coding region to that of MET25

Cited by 75 publications

References 28 publications

The Ras/cAMP-dependent Protein Kinase Signaling Pathway Regulates an Early Step of the Autophagy Process in Saccharomyces cerevisiae

The Ras/cAMP-dependent Protein Kinase Signaling Pathway Regulates an Early Step of the Autophagy Process in Saccharomyces cerevisiae

Site-selected Mutagenesis of a Conserved Nucleotide Binding HXGH Motif Located in the ATP Sulfurylase Domain of Human Bifunctional 3′-Phosphoadenosine 5′-Phosphosulfate Synthase

Specific Retrieval of the Exocytic SNARE Snc1p from Early Yeast Endosomes

Contact Info

Product

Resources

About