scholarly journals Acidic phospholipid species inhibit adenylate cyclase activity in rat liver plasma membranes

1986 ◽  
Vol 235 (1) ◽  
pp. 237-243 ◽  
Author(s):  
M D Houslay ◽  
L Needham ◽  
N J Dodd ◽  
A M Grey
Keyword(s):  
Phase Separation ◽  
Adenylate Cyclase ◽  
Plasma Membranes ◽  
Cyclase Activity ◽  
Adenylate Cyclase Activity ◽  
Lipid Phase ◽  
Liver Plasma Membranes ◽  
Native Liver ◽  
Rat Liver Plasma Membranes ◽  
Lipid Phase Separation

Incubation of rat liver plasma membranes with liposomes of dioleoyl phosphatidic acid (dioleoyl-PA) led to an inhibition of adenylate cyclase activity which was more pronounced when fluoride-stimulated activity was followed than when glucagon-stimulated activity was followed. If Mn2+ (5 mM) replaced low (5 mM) [Mg2+] in adenylate cyclase assays, or if high (20 mM) [Mg2+] were employed, then the perceived inhibitory effect of phosphatidic acid was markedly reduced when the fluoride-stimulated activity was followed but was enhanced for the glucagon-stimulated activity. The inhibition of adenylate cyclase activity observed correlated with the association of dioleoyl-PA with the plasma membranes. Adenylate cyclase activity in dioleoyl-PA-treated membranes, however, responded differently to changes in [Mg2+] than did the enzyme in native liver plasma membranes. Benzyl alcohol, which increases membrane fluidity, had similar stimulatory effects on the fluoride- and glucagon-stimulated adenylate cyclase activities in both native and dioleoyl-PA-treated membranes. Incubation of the plasma membranes with phosphatidylserine also led to similar inhibitory effects on adenylate cyclase and responses to Mg2+. Arrhenius plots of both glucagon- and fluoride-stimulated adenylate cyclase activity were different in dioleoyl-PA-treated plasma membranes, compared with native membranes, with a new ‘break’ occurring at around 16 degrees C, indicating that dioleoyl-PA had become incorporated into the bilayer. E.s.r. analysis of dioleoyl-PA-treated plasma membranes with a nitroxide-labelled fatty acid spin probe identified a new lipid phase separation occurring at around 16 degrees C with also a lipid phase separation occurring at around 28 degrees C as in native liver plasma membranes. It is suggested that acidic phospholipids inhibit adenylate cyclase by virtue of a direct headgroup specific interaction and that this perturbation may be centred at the level of regulation of this enzyme by the stimulatory guanine nucleotide regulatory protein NS.

scholarly journals Lysophosphatidylcholines can modulate the activity of the glucagon-stimulated adenylate cyclase from rat liver plasma membranes

1979 ◽  
Vol 178 (1) ◽  
pp. 217-221 ◽  
Author(s):  
M D Houslay ◽  
R W Palmer
Keyword(s):  
Adenylate Cyclase ◽  
Rat Liver ◽  
Plasma Membranes ◽  
Hormone Receptors ◽  
Cyclase Activity ◽  
Adenylate Cyclase Activity ◽  
Liver Plasma Membranes ◽  
Rat Liver Plasma Membranes ◽  
Increased Sensitivity

1. Synthetic lysophosphatidylcholines inhibit the glucagon-stimulated adenylate cyclase activity of rat liver plasma membranes at concentrations two to five times lower than those needed to inhibit the fluoride-stimulated activity. 2. Specific 125I-labelled glucagon binding to hormone receptors is inhibited at concentrations similar to those inhibiting the fluoride-stimulated activity. 3. At concentrations of lysophosphatidylcholines immediately below those causing inhibition, an activation of adenylate cyclase activity or hormone binding was observed. 4 These effects are essentially reversible. 5. We conclude that the increased sensitivity of glucagon-stimulated adenylate cyclase to inhibition may be due to the lysophosphatidylcholines interfering with the physical coupling between the hormone receptor and catalytic unit of adenylate cyclase. 6. We suggest that, in vivo, it is possible that lysophosphatidylcholines may modulate the activity of adenylate cyclase only when it is in the hormone-stimulated state.

scholarly journals Elevated membrane cholesterol concentrations inhibit glucagon-stimulated adenylate cyclase

1983 ◽  
Vol 210 (2) ◽  
pp. 437-449 ◽  
Author(s):  
A D Whetton ◽  
L M Gordon ◽  
M D Houslay
Keyword(s):  
Fatty Acid ◽  
Adenylate Cyclase ◽  
Spin Probe ◽  
Plasma Membranes ◽  
Cyclase Activity ◽  
Adenylate Cyclase Activity ◽  
Cholesterol Concentration ◽  
Lipid Phase ◽  
Lipid Order ◽  
Liver Plasma Membranes

A method was devised which increases the cholesterol concentration of rat liver plasma membranes by exchange from cholesterol-rich liposomes at low temperature (4 degrees C). When the cholesterol concentration of liver plasma membranes is increased, there is an increase in lipid order as detected by a decrease in mobility of an incorporated fatty acid spin probe. This is accompanied by an inhibition of adenylate cyclase activity. The various ligand-stimulated adenylate cyclase activities exhibit different sensitivities to inhibition by cholesterol, with inhibition of glucagon-stimulated greater than fluoride-stimulated greater than basal activity. The bilayer-fluidizing agent benzyl alcohol is able to reverse the inhibitory effect of cholesterol on adenylate cyclase activity in full. The thermostability of fluoride-stimulated cyclase is increased in the cholesterol-rich membranes. Elevated cholesterol concentrations abolish the lipid-phase separation occurring at 28 degrees C in native membranes as detected by an incorporated fatty acid spin probe. This causes Arrhenius plots of glucagon-stimulated adenylate cyclase activity to become linear, rather than exhibiting a break at 28 degrees C. It is suggested that the cholesterol contents of both halves of the bilayer are increased by the method used and that inhibition of adenylate cyclase ensues, owing to the increase in lipid order and promotion of protein-protein and specific cholesterol-phospholipid interactions.

scholarly journals Glucagon-stimulated adenylate cyclase detects a selective perturbation of the inner half of the liver plasma-membrane bilayer achieved by the local anaesthetic prilocaine

1980 ◽  
Vol 190 (1) ◽  
pp. 131-137 ◽  
Author(s):  
M D Houslay ◽  
I Dipple ◽  
S Rawal ◽  
R D Sauerheber ◽  
J A Esgate ◽  
...  
Keyword(s):  
Phase Separation ◽  
Fatty Acid ◽  
Adenylate Cyclase ◽  
Rat Liver ◽  
Spin Probe ◽  
Plasma Membranes ◽  
Lipid Phase ◽  
Rat Liver Plasma Membranes ◽  
Membrane Bound ◽  
Lipid Phase Separation

Prilocaine can increase the fluidity of rat liver plasma membranes, as indicated by a fatty acid spin-probe. This led to the activation of the membrane-bound fluoride-stimulated adenylate cyclase activity, but not the Lubrol-solubilized activity, suggesting that increased lipid fluidity can activate the enzyme. With increasing prilocaine concentrations above 10 mM, the membrane-bound fluoride-stimulated activity was progressively inhibited, even though bilayer fluidity continued to increase and the activity of the solubilized enzyme remained unaffected. Glucagon-stimulated adenylate cyclase was progressively inhibited by increasing prilocaine concentrations. Prilocaine (10 mM) had no effect on the lipid phase separation occurring at 28 degrees C and attributed to those lipids in the external half of the bilayer, as indicated by Arrhenius plots of both glucagon-stimulated adenylate cyclase activity and the order parameter of a fatty acid spin-probe. However, 10 mM-prilocaine induced a lipid phase separation at around 11 degrees C that was attributed to the lipids of the internal (cytosol-facing) half of the bilayer. It is suggested that prilocaine (10 mM) can selectively perturb the inner half of the bilayer of rat liver plasma membranes owing to its preferential interaction with the acidic phospholipids residing there.

scholarly journals The activity of glucagon-stimulated adenylate cyclase from rat liver plasma membranes is modulated by the fluidity of its lipid environment

1978 ◽  
Vol 174 (1) ◽  
pp. 179-190 ◽  
Author(s):  
I Dipple ◽  
M D Houslay
Keyword(s):  
Adenylate Cyclase ◽  
Benzyl Alcohol ◽  
Plasma Membranes ◽  
Break Point ◽  
Alcohol Concentration ◽  
Cyclase Activity ◽  
Adenylate Cyclase Activity ◽  
Lipid Phase ◽  
Glucagon Receptor ◽  
Arrhenius Plots

1. The local anaesthetic benzyl alcohol progressively activated glucagon-stimulated adenylate cyclase activity up to a maximum at 50 mM-benzyl alcohol. Further increases in benzyl alcohol concentration inhibited the activity. The fluoride-stimulated adenylate cyclase activity was similarly affected except for an inhibition of activity occurring at low benzyl alcohol concentrations (approx. 10 mM. 2. The fluoride-stimulated adenylate cyclase activity of a solubilized enzyme preparation was unaffected by any of the benzyl alcohol concentrations tested. 3. Increases in 3-phenylpropan-1-ol and 5-phenylpentan-1-ol concentrations progressively activated both the fluoride- and glucagon-stimulated adenylate cyclase activities up to a maximum, above which further increases in alcohol concentration inhibited the activities. 4. The ‘break’ points in Arrhenius plots of glucagon-stimulated adenylate cyclase activity in native plasma membranes, and in plasma membranes fused with synthetic dimyristoyl phosphatidylcholine so as to constitute 60% of the total lipid pool, were decreased by approx. 6 degrees C by addition of 40 mM-benzyl alcohol. This was accompanied by a fall in the associated activation energies. 6. Arrhenius plots of fluoride-stimulated adenylate cyclase activity in the presence and absence of 40 mM-benzyl alcohol were linear, although addition of benzyl alcohol caused a dramatic decrease in the associated activation energy of the reaction. 7. 5′-Nucleotidase activity was stimulated by benzyl alcohol, and the ‘break’ point in the Arrhenius plot of its activity was decreased by about 6 degrees C by addition of 40 mM-benzyl alcohol to the assay. 8. It is suggested that benzyl alcohol effects a fluidization of the bilayer, which is clearly demonstrated by its ability to lower the temperature of a lipid phase separation occurring at 28 degrees C in the outer half of the bilayer to around 22 degrees C. The increase in bilayer fluidity relieves a physical constraint on the membrane-bound adenylate cyclase, activating the enzyme. 9. The various inhibition phenomena are discussed in detail, together with the suggestion that the interaction between the uncoupled catalytic unit of adenylate cyclase and the lipids of the bilayer is altered on its physical coupling to the glucagon receptor.

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