Adrenaline and the regulation of acetyl-coenzyme A carboxylase in rat epididymal adipose tissue. Inactivation of the enzyme is associated with phosphorylation and can be reversed on dephosphorylation

Brownsey, Roger W.; Hughes, William A.; Denton, Richard M.

doi:10.1042/bj1840023

Cited by 98 publications

(24 citation statements)

References 27 publications

(23 reference statements)

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“…Since then this approach has been further developed by others including ourselves [112][113][114][115]. In particular, subcellular fractionation and specific immunoprecipitation have been employed to separate and identify a number of the major labelled phosphoproteins [18,67,[112][113][114][115][116][117]129]. Table 2 lists the principle alterations in protein phosphorylation which have been reported in fat cells exposed to insulin or adrenaline.…”

Section: Effects Of Insulin On Specific Protein Phosphorylation In Inmentioning

confidence: 99%

“…Techniques were therefore developed involving rapid immuno-isolation, trypsin digestion and separation of phosphopeptides which allow the relative level of phosphorylation of the sites to be assessed [67]. All three sites were shown to be phosphorylated in intact fat cells but no evidence for any change in the relative occupancy of the three sites was [116,117]. Following trypsin digestion the [3zP]-phosphopeptides were separated by two-dimensional analysis on thin layer cellulose plates [139].…”

Section: Effects Of Insulin On Specific Protein Phosphorylation In Inmentioning

confidence: 99%

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A partial view of the mechanism of insulin action

1981

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Insulin was discovered 60 years ago [1]; since then its structure has been determined by the pioneer work of Sanger [2] and by Hodgkin, Blundell and colleagues [3]. Moreover the principle effects of insulin on carbohydrate, fat and protein metabolism have been well recognised, at least ingeneral terms, for some 20 years [4][5][6][7], yet, we do not have a satisfactory description at the molecular level of how this hormone brings about its wide range of effects on target cells. This is certainly not due to any lack of attention to the problem by research workers. The literature on this subject is vast and, in the absence of any generally acceptable hypothesis, it also tends to be rather confused and certainly conflicting. Since it is quite unrealistic to survey the whole field, we make no apology for concentrating on certain aspects in this article.Insulin has both long and short term effects on the metabolism of its target cells. The long term effects involve changes in both general and specific protein synthesis and breakdown, while short term effects are those brought about solely through changes in the activity of pre-existing enzymes and membrane transporters. To a large extent, we will neglect all aspects of long term regulation including protein synthesis and amino acid metabolism. In fact, advances in these areas have been delayed by the lack of satisfactory preparations suitable for probing the mechanisms involved in the specific induction and repression of enzymes by insulin in vitro. This situation will probably change rapidly as a number of such preparations have been reported recently. Particularly promising are the 3T3-L1 preadipocyte cell line [8,9] and also the maintenance liver cell cultures from young rats in which glucokinase activity has been shown to be rapidly induced on exposure to insulin in the presence of glucose [10].In the rest of this review we will concentrate on those studies concerned with short term effects of insulin on carbohydrate and fat metabolism in the cells of liver, muscle and, particularly, adipose tissue. All these cells have specific insulin receptors on the outward face of the plasma membrane but we will not be considering the nature of these receptors nor their interaction with insulin nor the mechanisms which may govern the number of insulin receptors. These aspects have all been well covered in other recent reviews [11][12][13]. We shall address ourselves to the problem of the process or processes whereby alterations in the level of occupancy of the plasma membrane insulinreceptors lead to a wide range of intracellular changes. The assumption will be made that neither the whole insulin molecule nor part of it needsto enter the cells of its target tissue to bring about the effects of insulin. There seems little doubt that insulin bound to its receptor can be "internalized" into liver and probably other cells [14, 15], however, the process appears to be too slow to represent the means whereby insulin could bring about its short term effects. On the other hand, antibo...

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Section: Effects Of Insulin On Specific Protein Phosphorylation In Inmentioning

confidence: 99%

Section: Effects Of Insulin On Specific Protein Phosphorylation In Inmentioning

confidence: 99%

A partial view of the mechanism of insulin action

1981

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“…The concentration of protein kinase used for the experiment shown in Fig.2 is actually less than half the estimated concentrations of the protein kinase in vivo in rat liver and mammary glands [40]. The rate of phosphorylation in vitro is thus quite sufficient to account for effects of adrenaline or glucagon on the phosphorylation and inactivation of acetyl-CoA carboxylase in intact cells, which are observed within 10 min [7,8,10]. These results represent the first conclusive demonstration that the phosphorylation of acetyl-CoA carboxylase by cyclic-AMP-dependent protein kinase leads to a change in the kinetic properties of the enzyme.…”

Section: Discussionmentioning

confidence: 99%

“…This possibility received strong support when Hardie and Cohen [15] reported that highly purified acetyl-CoA carboxylase from rabbit mammary gland could be phosphorylated by cyclic-AMPdependent protein kinase and at least one other protein kinase, and Brownsey et al [16] detected the phosphorylation of the enzyme in intact fat cells. Subsequently it has been found that the labelling of acetyl-CoA carboxylase when rats are injected with inorganic [32P]phosphate or when isolated cells are incubated with inorganic [32P]phosphate, is stimulated by glucagon in the liver [lo], and by adrenaline in adipose tissue [7,8].…”

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confidence: 99%

Reversible Phosphorylation and Inactivation of Acetyl‐CoA Carboxylase from Lactating Rat Mammary Gland by Cyclic AMP‐Dependent Protein Kinase

Hardie

Guy

1980

European Journal of Biochemistry

110

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Acetyl-CoA carboxylase has been purified from lactating rat mammary gland using a combination of ammonium sulphate and poly(ethyleneglyco1) precipitations. The enzyme was purified from 35-70-fold with a yield of ovier SO%, the exact figures being difficult to estimate because of activation of the enzyme that occurs during the preparation. The preparation was homogeneous by the criterion of polyacrylamide gel electrophoresis in sodium dodecyl sulphate and had a single subunit of molecular weight 240000, containing 1.02 f 0.04 molecules of biotin and 3.1 +_ 1.7 molecules of alkali-labile phosphate per subunit. The purified enzyme was phosphorylated and inactivated rapidly when incubated in the presence of [Y-~'P]ATP and magnesium ions with the purified catalytic subunit of cyclic-AMP-dependent protein kinase from rabbit skeletal muscle. Both phosphorylation and inactivation are blocked by the heat-stable protein inhibitor of cyclic-AMP-dependent protein kinase, and can be reversed by incubation with purified protein phosphatase-1 from rabbit skeletal muscle. The inactivation by the protein kinase and reactivation by the protein phosphatase correlate with the near-stoichiometric phosphorylation and dephosphorylation of site(s) located in a single tryptic peptide. Phosphorylation does not affect the K,,, for substrates, but brings about a twofold decrease in V and a twofold increase in the apparent dissociation constant for the allosteric activator, citrate. We also present evidence that the activation of rabbit mammary acetyl-CoA carboxylase by protein phosphatase-1 described previously [Hardie and Cohen (1979) FEBS Lett. 103, 333-3381 is due to dephosphorylation at site(s) which are not phosphorylated by either cyclic-AMP-dependent protein kinase or acetyl-CoA carboxylase kinase-2.These results suggest that the rapid inactivation of acetyl-CoA carboxylase, and hence fatty acid synthesis, by adrenaline in adipose tissue, or glucagon in the liver, is due to phosphorylation of the enzyme by cyclic-AMP-dependent protein kinase.

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“…Phosphorylation of ACC within Intact Adipose Tissue-Incubation of adipose tissue in the absence of hormones promotes formation of minimally phosphorylated forms of ACC (41,51). This treatment enhanced ACC activation by glutamate determined at zero or low concentrations of citrate, even when microcystin LR was present (Fig.…”

Section: Effects Of Glutamate Depend On the Extent Of Acc Phosphorylamentioning

confidence: 99%

Bimodal Activation of Acetyl-CoA Carboxylase by Glutamate

Boone¹,

Chan²,

Kulpa

et al. 2000

Journal of Biological Chemistry

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Acetyl-CoA carboxylase (ACC) catalyzes the formation of malonyl-CoA, an essential substrate for fatty acid biosynthesis and a potent inhibitor of fatty acid oxidation. Here, we provide evidence that glutamate may be a physiologically relevant activator of ACC. Glutamate induced the activation of both major isoforms of ACC, prepared from rat liver, heart, or white adipose tissue. In agreement with previous studies, a type 2A protein phosphatase contributed to the effects of glutamate on ACC. However, the protein phosphatase inhibitor microcystin LR did not abolish the effects of glutamate on ACC activity. Moreover, glutamate directly activated purified preparations of ACC when protein phosphatase activity was excluded. Phosphatase-independent ACC activation by glutamate was also reflected by polymerization of the enzyme as judged by size-exclusion chromatography. The sensitivity of ACC to direct activation by glutamate was diminished by treatment in vitro with AMP-activated protein kinase or cAMP-dependent protein kinase or by ␤-adrenergic stimulation of intact adipose tissue. We conclude that glutamate, an abundant intracellular amino acid, induces ACC activation through complementary actions as a phosphatase activator and as a direct allosteric ligand for dephosphorylated ACC. This study supports the general hypothesis that amino acids fulfill important roles as signal molecules as well as intermediates in carbon and nitrogen metabolism.

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Adrenaline and the regulation of acetyl-coenzyme A carboxylase in rat epididymal adipose tissue. Inactivation of the enzyme is associated with phosphorylation and can be reversed on dephosphorylation

Cited by 98 publications

References 27 publications

A partial view of the mechanism of insulin action

A partial view of the mechanism of insulin action

Reversible Phosphorylation and Inactivation of Acetyl‐CoA Carboxylase from Lactating Rat Mammary Gland by Cyclic AMP‐Dependent Protein Kinase

Bimodal Activation of Acetyl-CoA Carboxylase by Glutamate

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