Regulation of carnitine palmitoyltransferase in vivo by glucagon and insulin

Abstract

Carnitine palmitoyltransferase (CPT total) activity and synthesis increase in states where the insulin/glucagon ratio is low, such as starvation and diabetes [Brady & Brady (1987) Biochem. J. 246, 641-646]. However, the effect of glucagon and insulin on CPT synthesis is unknown. The present experiments were designed to determine the effect of glucagon, cAMP [8-(chlorophenylthio) cyclic AMP], and insulin + cAMP on CPT transcription and mRNA amounts over time after injection. The CPT protein that was purified, used to generate antibody, and cloned in these studies was the 68 kDa mitochondrial protein described previously [Brady & Brady (1987) Biochem. J. 246, 641-646; Brady, Feng & Brady (1988) J. Nutr. 118, 1128-1136; Brady & Brady (1989) Diabetes 38, in the press]. Saline-injected control rats exhibited a 2-fold increase in hepatic CPT transcription rate and CPT mRNA over the 5 h experiment from 09:00 to 14:00 h. The effect was most probably due to the fasting state of the rats during the day. Glucagon injection caused an 8-fold increase in transcription rate by 90 min and a 4-fold increase in CPT mRNA by 90-120 min. The cAMP effect had reached a peak by the first time point taken (15 min). Transcription rate was increased 4-fold and CPT mRNA was increased 3-fold at this time. The combination of cAMP + insulin injection did not produce any significant increase in transcription rate or CPT mRNA over the saline-injected controls. CPT mRNA and transcription rate showed a clear dose-response to glucagon injection from 0 to 150 micrograms/100 g body wt. Total CPT activity and immunoreactive CPT were not increased during these experiments. The data indicate that glucagon and insulin interact in control of transcription rate and amount of CPT mRNA, but that increases in CPT immunoreactive protein and activity are temporally delayed. This lag probably relates to the half-life of the CPT protein in vivo, which has been estimated as 2-7 days.

Enhanced Potential for Oxidative Stress in Hyperinsulinemic Rats: Imbalance Between Hepatic Peroxisomal Hydrogen Peroxide Production and Decomposition Due to Hyperinsulinemia

“…In agreement with resuits obtained in this study, insulin produced a pronounced inhibitory effect on the transcription of mRNA of peroxisomal 1oxidizing enzymes in the liver [16]. Furthermore, isulin supplementation depressed both content as well as transcription Glucose of carnitine palmitoyltransferase mRNA [19]. In this study, the decrease in catalase activity is most likely directly related to the observed inhibitory effect of insulin on the synthesis of the catalase protein (Fig.…”

Turnover of carnitine palmitoyltransferase mRNA and protein in H4IIE cells. Effect of cyclic AMP and insulin

“…The CPT was detected by autoradiography and the band cut out and counted for radioactivity by liquid scintillation. in CPT activity or immunoreactive protein (Brady & Brady, 1989b), and (2) CPT-protein t2 was 2-7.5 days as measured in vivo (Ozasa et al, 1983;Brady & Brady, 1987), but CPT-mRNA t1 in vivo was 6-8 h. Our main question, then, was 'what is the relationship between CPT-mRNA turnover and CPT-protein turnover? '.…”

“…In BB Wistar spontaneously diabetic rats, hepatic CPT activity, CPT mRNA and transcription rates are also increased 2-6-fold, and return to control values on insulin administration (Brady & Brady, 1989a). When glucagon or cyclic AMP is administered in vivo, there is a 3-5-fold increase in hepatic CPT transcription rate and CPT mRNA within 1 h, but no increase in CPT activity or immunoreactive protein (Brady & Brady, 1989b). These results suggest that in vivo the halflife (ti) of CPT mRNA and of CPT protein may be quite different.…”

Activity of carnitine palmitoyltransferase in mitochondrial outer membranes and peroxisomes in digitonin-permeabilized hepatocytes. Selective modulation of mitochondrial enzyme activity by okadaic acid

“…The okadaic acid-induced stimulation of hepatic mitochondrial palmitate oxidation seems to be due (at least in part) to profound changes in the kinetic and regulatory properties of CPTO, namely increased specific activity, decreased sensitivity to inhibition by malonyl-CoA and decreased affinity for palmitoyl-CoA substrate. Although the exhaustive molecular characterization of CPTi has allowed the study of the mechanisms involved in the long-term regulation of this enzyme protein [8,29,35], the molecular processes underlying the alterations in the kinetic and regulatory properties of hepatic CPTO are still unknown [2,3,9,10]. Very recently a polyclonal antibody against rat liver CPTO has been obtained, and the levels of immunoreactive CPTO protein have been shown to increase several-fold under different ketotic states [36].…”