The lipid composition of plasma membrane and mitochondrial fractions from epididymal adipocytes of cold-acclimated rats

Cherqui, Gisèle; Cadot, M; Senault, Claude; Portet, R

doi:10.1016/0005-2736(89)90008-4

Cited by 9 publications

(1 citation statement)

References 39 publications

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“…(2) Liver plasma membranes were judged to be more appropriate than adipocyte plasma membranes for testing the effect of insulin on membrane proteolysis. This choice allowed us to obviate a problem inherent in the preparation of adipocyte plasma membranes, that is the achievement of fat-cell disruption along with subsequent centrifugation of the resulting homogenate at a temperature of 20°C (Avruch & Wallach, 1971;Cherqui et al, 1979).…”

Section: Discussionmentioning

confidence: 99%

Further evidence for the involvement of a membrane proteolytic step in insulin action

Cherqui¹,

Caron²,

Capeau³

et al. 1985

Biochemical Journal

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The hypothesis that insulin action involves a membrane proteolytic step was further explored, by using isolated rat adipocytes and liver plasma membranes. (1) The maximal insulin stimulation of 2-deoxyglucose transport and lipogenesis in fat-cells was selectively inhibited (73-88%) by N alpha-p-tosyl-L-lysine chloromethyl ketone (Tos-Lys-CH2Cl; active-site inhibitor of trypsin; 30-125 microM), p-nitrophenyl p'-guanidinobenzoate (active-site inhibitor of serine proteinases; 30-125 microM) and p-tosyl-L-arginine methyl ester (arginine ester substrate analogue of proteinases; 1-2 mM), under conditions where neither the basal rate of each metabolic process nor insulin binding nor cellular ATP content were affected. In contrast, N-acetyl-L-alanyl-L-alanyl-L-alanine methyl ester (alanine ester substrate analogue of proteinases; 1-2 mM) was ineffective. (2) Endoproteinase Arg-C (0.25-40 micrograms/ml) exerted dose-dependent insulin-like effects on both 2-deoxyglucose transport and lipogenesis in fat-cells, whereas endoproteinase Lys-C (5-100 micrograms/ml) was ineffective. The maximal activation by endoproteinase Arg-C of both processes (200 and 177% of control values respectively) was shown to occur under conditions where membrane integrity (assessed by measurement of lactate dehydrogenase leakage and passive glucose diffusion) was preserved. This effect was inhibited by Tos-Lys-CH2Cl (125 microM) and was not additive with the maximal insulin effect. (3) Insulin (1-100 ng/ml) produced a dose-dependent increase in the trichloroacetic acid-soluble 125I radioactivity released after a 30 min incubation at 37 degrees C of 125I-labelled liver plasma membranes, but was ineffective on 125I-labelled bovine serum albumin. Insulin effects on both radio-labelled proteins were reproduced by wheat-germ agglutinin (20 micrograms/ml), an insulin mimicker shown to act through the insulin receptor. These data provide further evidence for the hypothesis that insulin bioeffects involve the activation of a membrane serine proteinase with arginine specificity.

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Section: Discussionmentioning

confidence: 99%