scholarly journals Changes in gene expression in the Ras/adenylate cyclase system of Saccharomyces cerevisiae: correlation with cAMP levels and growth arrest.

1993 ◽  
Vol 4 (7) ◽  
pp. 757-765 ◽  
Author(s):  
M Russell ◽  
J Bradshaw-Rouse ◽  
D Markwardt ◽  
W Heideman
Keyword(s):  
Gene Expression ◽  
Saccharomyces Cerevisiae ◽  
Adenylate Cyclase ◽  
Oxidative Metabolism ◽  
Growth Arrest ◽  
Intracellular Camp ◽  
Adenylate Cyclase System ◽  
Diauxic Shift ◽  
Yeast Saccharomyces Cerevisiae ◽  
Camp Levels

Levels of cyclic 3',5'-cyclic monophosphate (cAMP) play an important role in the decision to enter the mitotic cycle in the yeast, Saccharomyces cerevisiae. In addition to growth arrest at stationary phase, S. cerevisiae transiently arrest growth as they shift from fermentative to oxidative metabolism (the diauxic shift). Experiments examining the role of cAMP in growth arrest at the diauxic shift show the following: 1) yeast lower cAMP levels as they exhaust their glucose supply and shift to oxidative metabolism of ethanol, 2) a reduction in cAMP is essential for traversing the diauxic shift, 3) the decrease in adenylate cyclase activity is associated with a decrease in the expression of CYR1 and CDC25, two positive regulators of cAMP levels and an increase in the expression of IRA1 and IRA2, two negative regulators of intracellular cAMP, 4) mutants carrying disruptions in IRA1 and IRA2 were unable to arrest cell division at the diauxic shift and were unable to progress into the oxidative phase of growth. These results indicate that changes cAMP levels are important in regulation of growth arrest at the diauxic shift and that changes in gene expression plays a role in the regulation of the Ras/adenylate cyclase system.

scholarly journals High cAMP Levels Antagonize the Reprogramming of Gene Expression that Occurs at the Diauxic Shift in Saccharomyces Cerevisiae

Microbiology ◽  
1996 ◽  
Vol 142 (3) ◽  
pp. 459-467 ◽  
Author(s):  
E. Boy-Marcotte ◽  
D. Tadi ◽  
M. Perrot ◽  
H. Boucherie ◽  
M. Jacquet
Keyword(s):  
Gene Expression ◽  
Saccharomyces Cerevisiae ◽  
Diauxic Shift ◽  
Camp Levels

Adenylate cyclase in Saccharomyces cerevisiae is a peripheral membrane protein

1990 ◽  
Vol 10 (8) ◽  
pp. 3873-3883
Author(s):  
M R Mitts ◽  
D B Grant ◽  
W Heideman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Membrane Protein ◽  
Adenylate Cyclase ◽  
Adenylate Cyclase System ◽  
Hydrodynamic Properties ◽  
Yeast Saccharomyces Cerevisiae ◽  
Peripheral Membrane Protein ◽  
Mammalian Enzyme ◽  
Regulatory Significance ◽  
Stimulation Of

The adenylate cyclase system of the yeast Saccharomyces cerevisiae contains the CYR1 polypeptide, responsible for catalyzing formation of cyclic AMP (cAMP) from ATP, and two RAS polypeptides, which mediate stimulation of cAMP synthesis of guanine nucleotides. By analogy to the mammalian enzyme, models of yeast adenylate cyclase have depicted the enzyme as a membrane protein. We have concluded that adenylate cyclase is only peripherally bound to the yeast membrane, based on the following criteria: (i) substantial activity was found in cytoplasmic fractions; (ii) activity was released from membranes by the addition of 0.5 M NaCl; (iii) in the presence of 0.5 M NaCl, activity in detergent extracts had hydrodynamic properties identical to those of cytosolic or NaCl-extracted enzyme; (iv) antibodies to yeast adenylate cyclase identified a full-length adenylate cyclase in both membrane and cytosol fractions; and (v) activity from both cytosolic fractions and NaCl extracts could be functionally reconstituted into membranes lacking adenylate cyclase activity. The binding of adenylate cyclase to the membrane may have regulatory significance; the fraction of activity associated with the membrane increased as cultures approached stationary phase. In addition, binding of adenylate cyclase to membranes appeared to be inhibited by cAMP. These results indicate the existence of a protein anchoring adenylate cyclase to the membrane. The identity of this protein remains unknown.

scholarly journals Adenylate cyclase in Saccharomyces cerevisiae is a peripheral membrane protein.

1990 ◽  
Vol 10 (8) ◽  
pp. 3873-3883 ◽  
Author(s):  
M R Mitts ◽  
D B Grant ◽  
W Heideman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Membrane Protein ◽  
Adenylate Cyclase ◽  
Adenylate Cyclase System ◽  
Hydrodynamic Properties ◽  
Yeast Saccharomyces Cerevisiae ◽  
Peripheral Membrane Protein ◽  
Mammalian Enzyme ◽  
Regulatory Significance ◽  
Stimulation Of

The adenylate cyclase system of the yeast Saccharomyces cerevisiae contains the CYR1 polypeptide, responsible for catalyzing formation of cyclic AMP (cAMP) from ATP, and two RAS polypeptides, which mediate stimulation of cAMP synthesis of guanine nucleotides. By analogy to the mammalian enzyme, models of yeast adenylate cyclase have depicted the enzyme as a membrane protein. We have concluded that adenylate cyclase is only peripherally bound to the yeast membrane, based on the following criteria: (i) substantial activity was found in cytoplasmic fractions; (ii) activity was released from membranes by the addition of 0.5 M NaCl; (iii) in the presence of 0.5 M NaCl, activity in detergent extracts had hydrodynamic properties identical to those of cytosolic or NaCl-extracted enzyme; (iv) antibodies to yeast adenylate cyclase identified a full-length adenylate cyclase in both membrane and cytosol fractions; and (v) activity from both cytosolic fractions and NaCl extracts could be functionally reconstituted into membranes lacking adenylate cyclase activity. The binding of adenylate cyclase to the membrane may have regulatory significance; the fraction of activity associated with the membrane increased as cultures approached stationary phase. In addition, binding of adenylate cyclase to membranes appeared to be inhibited by cAMP. These results indicate the existence of a protein anchoring adenylate cyclase to the membrane. The identity of this protein remains unknown.

scholarly journals Interactions between adenylate cyclase and the yeast GTPase-activating protein IRA1.

1991 ◽  
Vol 11 (9) ◽  
pp. 4591-4598 ◽  
Author(s):  
M R Mitts ◽  
J Bradshaw-Rouse ◽  
W Heideman
Keyword(s):  
Cyclic Amp ◽  
Adenylate Cyclase ◽  
Adenylate Cyclase System ◽  
Gtpase Activating Protein ◽  
Yeast Saccharomyces Cerevisiae ◽  
Strong Candidate ◽  
Sepharose 4B ◽  
Stimulation Of ◽  
Cyclic Amp Synthesis

The adenylate cyclase system of the yeast Saccharomyces cerevisiae contains many proteins, including the CYR1 polypeptide, which is responsible for catalyzing the formation of cyclic AMP from ATP, RAS1 and RAS2 polypeptides, which mediate stimulation of cyclic AMP synthesis by guanine nucleotides, and the yeast GTPase-activating protein analog IRA1. We have previously reported that adenylate cyclase is only peripherally bound to the yeast membrane. We have concluded that IRA1 is a strong candidate for a protein involved in anchoring adenylate cyclase to the membrane. We base this conclusion on the following criteria: (i) a disruption of the IRA1 gene produced a mutant with very low membrane-associated levels of adenylate cyclase activity, (ii) membranes made from these mutants were incapable of binding adenylate cyclase in vitro, (iii) IRA1 antibodies inhibit binding of adenylate cyclase to the membrane, and (iv) IRA1 and adenylate cyclase comigrate on Sepharose 4B.

Properties and Possible Functions of the Adenylate Cyclase in Plasma Membranes of Saccharomyces cerevisiae

1982 ◽  
Vol 2 (12) ◽  
pp. 1481-1491
Author(s):  
Patrick K. Jaynes ◽  
James P. McDonough ◽  
Henry R. Mahler
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Protein Kinase ◽  
Adenylate Cyclase ◽  
Plasma Membranes ◽  
Dissociation Constants ◽  
Guanine Nucleotides ◽  
Adenylate Cyclase System ◽  
Regulatory Subunits ◽  
Molecular Weights ◽  
Essential Components

We have examined the possible role of adenosine 3′,5′-phosphate (cAMP) in functions associated with the plasma membranes of Saccharomyces cerevisiae. Purified membranes from this source contained an adenylate cyclase which was insensitive to activation by fluoride or guanine nucleotides, only weakly responsive to changes of carbon source in the growth medium, and strongly stimulated by vanadate. They also contained at least two classes of receptor proteins for guanine nucleotides (as measured by binding of labeled 5′-guanylyl methylene diphosphate) with apparent dissociation constants equal to 1.0 × 10 −7 and 3 × 10 −6 M, a protein kinase capable of phosphorylating added histones, the activity of which was stimulated by cAMP, and cAMP receptors that may function as regulatory subunits for this kinase. Membrane proteins were also susceptible to phosphorylation by endogenous kinase(s), with polypeptides of apparent molecular weights equal to 160 × 10 3 , 135 × 10 3 , 114 × 10 3 , and 58 × 10 3 as the major targets. Of these, the 114,000-molecular-weight polypeptide was probably identical to the proton-translocating ATPase of the membranes. However, the cAMP-dependent protein kinase did not appear to be involved in these reactions. Intact ( rho + or rho 0 ) cells responded to dissipation of the proton electrochemical gradient across their plasma membranes by rapid and transient changes in their intracellular level of cAMP, as suggested earlier (J. M. Trevillyan and M. L. Pall, J. Bacteriol., 138 :397-403, 1979). Thus, although yeast plasma membranes contain all the essential components of a stimulus-responsive adenylate cyclase system, the precise nature of the coupling device and the targets involved remain to be established.

scholarly journals PGE2-mediated inhibition of T cell p59fynis independent of cAMP

1999 ◽  
Vol 277 (2) ◽  
pp. C302-C309 ◽  
Author(s):  
Mashkoor A. Choudhry ◽  
Zulfiqar Ahmed ◽  
Mohammed M. Sayeed
Keyword(s):  
T Cells ◽  
T Cell ◽  
Adenylate Cyclase ◽  
Protein Tyrosine Kinase ◽  
Kinase Activity ◽  
Prostaglandin E ◽  
Intracellular Camp ◽  
Sprague Dawley ◽  
Cell Functions ◽  
Camp Levels

We recently observed that prostaglandin E2(PGE2)-mediated suppression of T cell functions could result from an attenuation of p59fynprotein tyrosine kinase activity. The present study evaluated the effects of an adenylate cyclase agonist (forskolin) and antagonist (SQ-22536), as well as those of cAMP analogues (dibutyryl cAMP and 8-bromo- cAMP), on T cell p59fynkinase activity. The study allowed us to assess whether PGE2-mediated activation of adenylate cyclase by itself or the elevation in intracellular cAMP levels is an integral event in the modulation of anti-CD3-linked p59fynactivation in T cells. The experiments were carried out with splenic T cells from male Sprague-Dawley rats. A 30–50% suppression in the autophosphorylation and the kinase activity of p59fynin T cells incubated with PGE2or forskolin was observed. Pretreatment of T cells with SQ-22536 prevented significant PGE2-mediated inhibition of T cell p59fynkinase activity. In contrast, no change in p59fynautophosphorylation and kinase activity in T cells treated with cAMP analogues was observed. These data suggest that PGE2-mediated suppression of p59fynautophosphorylation and kinase activity in T cells is dependent on the activation of adenylate cyclase and independent of the elevation in cAMP levels.

Positive control of sporulation-specific genes by the IME1 and IME2 products in Saccharomyces cerevisiae

1990 ◽  
Vol 10 (5) ◽  
pp. 2104-2110
Author(s):  
A P Mitchell ◽  
S E Driscoll ◽  
H E Smith
Keyword(s):  
Gene Expression ◽  
Saccharomyces Cerevisiae ◽  
Meiotic Recombination ◽  
Positive Control ◽  
Spore Formation ◽  
Specific Gene ◽  
Heterologous Promoter ◽  
Yeast Saccharomyces Cerevisiae ◽  
Specific Gene Expression ◽  
Rapid Induction

In the yeast Saccharomyces cerevisiae, meiosis and spore formation require the induction of sporulation-specific genes. Two genes are thought to activate the sporulation program: IME1 and IME2 (inducer of meiosis). Both genes are induced upon entry into meiosis, and IME1 is required for IME2 expression. We report here that IME1 is essential for expression of four sporulation-specific genes. In contrast, IME2 is not absolutely essential for expression of the sporulation-specific genes, but contributes to their rapid induction. Expression of IME2 from a heterologous promoter permits the expression of these sporulation-specific genes, meiotic recombination, and spore formation in the absence of IME1. We propose that the IME1 and IME2 products can each activate sporulation-specific genes independently. In addition, the IME1 product stimulates sporulation-specific gene expression indirectly through activation of IME2 expression.

scholarly journals Role of IME1 expression in regulation of meiosis in Saccharomyces cerevisiae.

1990 ◽  
Vol 10 (12) ◽  
pp. 6103-6113 ◽  
Author(s):  
H E Smith ◽  
S S Su ◽  
L Neigeborn ◽  
S E Driscoll ◽  
A P Mitchell
Keyword(s):  
Gene Expression ◽  
Saccharomyces Cerevisiae ◽  
Amino Acid ◽  
Specific Gene ◽  
Alpha Cells ◽  
Cell Type ◽  
Yeast Saccharomyces Cerevisiae ◽  
Specific Gene Expression ◽  
Gal1 Promoter ◽  
Acid Protein

Two signals are required for meiosis and spore formation in the yeast Saccharomyces cerevisiae: starvation and the MAT products a1 and alpha 2, which determine the a/alpha cell type. These signals lead to increased expression of the IME1 (inducer of meiosis) gene, which is required for sporulation and sporulation-specific gene expression. We report here the sequence of the IME1 gene and the consequences of IME1 expression from the GAL1 promoter. The deduced IME1 product is a 360-amino-acid protein with a tyrosine-rich C-terminal region. Expression of PGAL1-IME1 in vegetative a/alpha cells led to moderate accumulation of four early sporulation-specific transcripts (IME2, SPO11, SPO13, and HOP1); the transcripts accumulated 3- to 10-fold more after starvation. Two sporulation-specific transcripts normally expressed later (SPS1 and SPS2) did not accumulate until PGAL1-IME1 strains were starved, and the intact IME1 gene was not activated by PGAL1-IME1 expression. In a or alpha cells, which lack alpha 2 or a1, expression of PGAL1-IME1 led to the same pattern of IME2 and SPO13 expression as in a/alpha cells, as measured with ime2::lacZ and spo13::lacZ fusions. Thus, in wild-type strains, the increased expression of IME1 in starved a/alpha cells can account entirely for cell type control, but only partially for nutritional control, of early sporulation-specific gene expression. PGAL1-IME1 expression did not cause growing cells to sporulate but permitted efficient sporulation of amino acid-limited cells, which otherwise sporulated poorly. We suggest that IME1 acts primarily as a positive regulator of early sporulation-specific genes and that growth arrest is an independent prerequisite for execution of the sporulation program.

cAMP metabolism and lipolysis in brown adipocytes of hamsters consuming a cafeteria diet

1985 ◽  
Vol 248 (2) ◽  
pp. E224-E229
Author(s):  
R. J. Schimmel ◽  
L. McCarthy ◽  
K. K. McMahon
Keyword(s):  
Adenylate Cyclase ◽  
Sympathetic Activity ◽  
Brown Fat ◽  
Adenylate Cyclase System ◽  
Brown Adipocytes ◽  
Beta Adrenergic ◽  
Fat Depots ◽  
Brown Adipose ◽  
Camp Levels ◽  
In Cells

Feeding animals cafeteria diets causes increased sympathetic activity to brown adipose tissue and this is believed to be responsible for the concomitant activation of thermogenesis. Because chronic catecholamine stimulation in other systems leads to a desensitization of beta-adrenergic receptors, we examined lipolysis and cAMP production in brown adipocytes of hamsters eating cafeteria diets for evidence of diminished beta-adrenergic responses. Basal cAMP levels were similar in chow- and cafeteria-fed hamsters. However, adipocytes from overfed animals formed less cAMP in response to isoproterenol than those of control animals. Isoproterenol-stimulated adenylate cyclase activity was similarly decreased in membrane preparations from cafeteria-fed hamsters. However, when the diterpene forskolin was used, equal amounts of cAMP were formed in cells and membrane preparations from control and overfed animals. In contrast to the reduced responses of the cAMP system to isoproterenol stimulation observed in overfed hamsters, isoproterenol-stimulated lipolysis was greater in cells from overfed animals than in cells from control animals. These results are consistent with a desensitization of the adenylate cyclase system in brown adipocytes occurring during chronic hyperphagia. Because eating cafeteria diets has been reported to increase sympathetic activity to brown fat depots, the apparent desensitization of brown adipocytes observed in this study may result from a persistent stimulation of the brown fat with norepinephrine in vivo. Our data also suggest the existence of mechanisms that preserve lipolysis in the face of low cAMP levels.

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