A purine-related metabolite negatively regulates fixNOQP expression in Sinorhizobium meliloti by modulation of fixK expression.
(25/623)
5-aminoimidazole-4-carboxamide nucleotide (AICAR) is a negative effector of cytochrome terminal oxidase cbb3 production in Rhizobium etli. In this work, the effect of AICAriboside (AICAr), the precursor of AICAR on the expression of the Sinorhizobium meliloti fixNOQP operon encoding the symbiotic terminal oxidase cbb3, was analyzed. AICAr reduced the microaerobic induction levels of fixN-lacZ and fixT-lacZ gene fusions 18- and seven-fold respectively, and both genes were activated by the transcriptional activator FixK. A fixK-lacZ fusion presented 14-fold-reduced induction levels in microaerobic cell cultures in the presence of AICAr. AICAr also reduced three-fold the microaerobic expression levels of the nifA-lacZ fusion, whose expression as well as that of fixK is controlled by the two-component system FixL-FixJ. In contrast, AICAr had no effect on the expression levels of a hemA-lacZ fusion. These data suggest that AICAr prevents fixNOQP induction by the inhibition of fixK transcription. (+info)
Effect of 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside infusion on in vivo glucose and lipid metabolism in lean and obese Zucker rats.
(26/623)
Activation of AMP-activated protein kinase (AMPK) with 5-aminoimidazole-4-carboxamide-1-beta-D-ribofurano-side (AICAR) increases glucose transport in skeletal muscle via an insulin-independent pathway. To examine the effects of AMPK activation on skeletal muscle glucose transport activity and whole-body carbohydrate and lipid metabolism in an insulin-resistant rat model, awake obese Zuckerfa/fa rats (n = 26) and their lean (n = 23) littermates were infused for 90 min with AICAR, insulin, or saline. The insulin infusion rate (4 mU.kg(-1).min(-1)) was selected to match the glucose requirements during AICAR (bolus, 100 mg/kg; constant, 10 mg.kg(-1).min(-1)) isoglycemic clamps in the lean rats. The effects of these identical AICAR and insulin infusion rates were then examined in the obese Zucker rats. AICAR infusion increased muscle AMPK activity more than fivefold (P < 0.01 vs. control and insulin) in both lean and obese rats. Plasma triglycerides, fatty acid concentrations, and glycerol turnover, as assessed by [2-13C]glycerol, were all decreased in both lean and obese rats infused with AICAR (P < 0.05 vs. basal), whereas insulin had no effect on these parameters in the obese rats. Endogenous glucose production rates, measured by [U-13C]glucose, were suppressed by >50% during AICAR and insulin infusions in both lean and obese rats (P < 0.05 vs. basal). In lean rats, rates of whole-body glucose disposal increased by more than two-fold (P < 0.05 vs. basal) during both AICAR and insulin infusion; [3H]2-deoxy-D-glucose transport activity increased to a similar extent, by >2.2-fold (both P < 0.05 vs. control), in both soleus and red gastrocnemius muscles of lean rats infused with either AICAR or insulin. In the obese Zucker rats, neither AICAR nor insulin stimulated whole-body glucose disposal or soleus muscle glucose transport activity. However, AICAR increased glucose transport activity by approximately 2.4-fold (P < 0.05 vs. control) in the red gastrocnemius from obese rats, whereas insulin had no effect. In summary, acute infusion of AICAR in an insulin-resistant rat model activates skeletal muscle AMPK and increases glucose transport activity in red gastrocnemius muscle while suppressing endogenous glucose production and lipolysis. Because type 2 diabetes is characterized by diminished rates of insulin-stimulated glucose uptake as well as increased basal rates of endogenous glucose production and lipolysis, these results suggest that AICAR-related compounds may represent a new class of antidiabetic agents. (+info)
Acute regulation of fatty acid oxidation and amp-activated protein kinase in human umbilical vein endothelial cells.
(27/623)
It is generally accepted that endothelial cells generate most of their ATP by anaerobic glycolysis and that very little ATP is derived from the oxidation of fatty acids or glucose. Previously, we have reported that, in cultured human umbilical vein endothelial cells (HUVECs), activation of AMP-activated protein kinase (AMPK) by the cell-permeable activator 5-aminoimidazole-4-carboximide riboside (AICAR) is associated with an increase in the oxidation of (3)H-palmitate. In the present study, experiments carried out with cultured HUVECs revealed the following: (1) AICAR-induced increases in palmitate oxidation during a 2-hour incubation are associated with a decrease in the concentration of malonyl coenzyme A (CoA) (an inhibitor of carnitine palmitoyl transferase 1), which temporally parallels the increase in AMPK activity and a decrease in the activity of acetyl CoA carboxylase (ACC). (2) AICAR does not stimulate either palmitate oxidation when carnitine is omitted from the medium or oxidation of the medium-chain fatty acid octanoate. (3) When intracellular lipid pools are prelabeled with (3)H-palmitate, the measured rate of palmitate oxidation is 3-fold higher, and in the presence of AICAR, it accounts for nearly 40% of calculated ATP generation. (4) Incubation of HUVECs in a glucose-free medium for 2 hours causes the same changes in AMPK, ACC, malonyl CoA, and palmitate oxidation as does AICAR. (5) Under all conditions studied, the contribution of glucose oxidation to ATP production is minimal. The results indicate that the AMPK-ACC-malonyl CoA-carnitine palmitoyl transferase 1 mechanism plays a key role in the physiological regulation of fatty acid oxidation in HUVECs. They also indicate that HUVECs oxidize fatty acids from both intracellular and extracellular sources, and that when this is taken into account, fatty acids can be a major substrate for ATP generation. Finally, they suggest that AMPK is likely to be a major factor in modulating the response of the endothelium to stresses that alter its energy state. (+info)
Hepatocyte nuclear factor-4alpha involved in type 1 maturity-onset diabetes of the young is a novel target of AMP-activated protein kinase.
(28/623)
Mutations in the HNF4alpha gene are responsible for type 1 maturity-onset diabetes of the young (MODY1), which is characterized by a defect in insulin secretion. Hepatocyte nuclear factor (HNF)-4alpha is a transcription factor that plays a critical role in the transcriptional regulation of genes involved in glucose metabolism in both hepatocytes and pancreatic beta-cells. Recent evidence has implicated AMP-activated protein kinase (AMPK) in the modulation of both insulin secretion by pancreatic beta-cells and the control of glucose-dependent gene expression in both hepatocytes and beta-cells. Therefore, the question could be raised as to whether AMPK plays a role in these processes by modulating HNF-4alpha function. In this study, we show that activation of AMPK by 5-amino-4-imidazolecarboxamide riboside (AICAR) in hepatocytes greatly diminished HNF-4alpha protein levels and consequently downregulates the expression of HNF-4alpha target genes. Quantitative evaluation of HNF-4alpha target gene expression revealed diminished mRNA levels for HNF-1alpha, GLUT2, L-type pyruvate kinase, aldolase B, apolipoprotein (apo)-B, and apoCIII. Our data clearly demonstrate that the MODY1/HNF-4alpha transcription factor is a novel target of AMPK in hepatocytes. Accordingly, it can be suggested that in pancreatic beta-cells, AMPK also acts by decreasing HNF-4alpha protein level, and therefore insulin secretion. Hence, the possible role of AMPK in the physiopathology of type 2 diabetes should be considered. (+info)
Activation of acetyl-CoA carboxylase by a glutamate- and magnesium-sensitive protein phosphatase in the islet beta-cell.
(29/623)
Acetyl-CoA carboxylase (ACC) catalyzes the formation of malonyl-CoA, a precursor in the biosynthesis of long-chain fatty acids, which have been implicated in physiological insulin secretion. The catalytic function of ACC is regulated by phosphorylation (inactive)-dephosphorylation (active). In this study we investigated whether similar regulatory mechanisms exist for ACC in the pancreatic islet beta-cell. ACC was quantitated in normal rat islets, human islets, and clonal beta-cells (HIT-15 or INS-1) using a [(14)C]bicarbonate fixation assay. In the beta-cell lysates, ACC was stimulated by magnesium in a concentration-dependent manner. Of all the dicarboxylic acids tested, only glutamate, albeit ineffective by itself, significantly potentiated magnesium-activated ACC in a concentration-dependent manner. ACC stimulation by glutamate and magnesium was maximally demonstrable in the cytosolic fraction; it was markedly reduced by okadaic acid (OKA) in concentrations (<50 nmol/l) that inhibited protein phosphatase 2A (PP2A). Furthermore, pretreatment of the cytosolic fraction with anti-PP2A serum attenuated the glutamate- and magnesium-mediated activation of ACC, thereby suggesting that ACC may be regulated by an OKA-sensitive PP2A-like enzyme. Streptavidin-agarose chromatography studies have indicated that glutamate- and magnesium-mediated effects on ACC are attributable to activation of ACC's dephosphorylation; this suggests that the stimulatory effects of glutamate and magnesium on ACC might involve activation of an OKA-sensitive PP2A-like enzyme that dephosphorylates and activates ACC. In our study, 5-amino-imidazolecarboxamide (AICA) riboside, a stimulator of AMP kinase, significantly inhibited glucose-mediated activation of ACC and insulin secretion from isolated beta-cells. Together, our data provide evidence for a unique regulatory mechanism for the activation of ACC in the pancreatic beta-cell, leading to the generation of physiological signals that may be relevant for physiological insulin secretion. (+info)
Regulation of fatty acid oxidation and glucose metabolism in rat soleus muscle: effects of AICAR.
(30/623)
Previous studies have shown that 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR), a cell-permeable activator of AMP-activated protein kinase, increases the rate of fatty acid oxidation in skeletal muscle of fed rats. The present study investigated the mechanism by which this occurs and, in particular, whether changes in the activity of malonyl-CoA decarboxylase (MCD) and the beta-isoform of acetyl-CoA carboxylase (ACC beta) are involved. In addition, the relationship between changes in fatty acid oxidation induced by AICAR and its effects on glucose uptake and metabolism was examined. In incubated soleus muscles isolated from fed rats, AICAR (2 mM) increased fatty acid oxidation (90%) and decreased ACC beta activity (40%) and malonyl-CoA concentration (50%); however, MCD activity was not significantly altered. In soleus muscles from overnight-fasted rats, AICAR decreased ACC beta activity (40%), as it did in fed rats; however, it had no effect on the already high rate of fatty acid oxidation or the low malonyl-CoA concentration. In keeping with its effect on fatty acid oxidation, AICAR decreased glucose oxidation by 44% in fed rats but did not decrease glucose oxidation in fasted rats. It had no effect on glucose oxidation when fatty acid oxidation was inhibited by 2-bromopalmitate. Surprisingly, AICAR did not significantly increase glucose uptake or assayable AMP-activated protein kinase activity in incubated soleus muscles from fed or fasted rats. These results indicate that, in incubated rat soleus muscle, 1) AICAR does not activate MCD or stimulate glucose uptake as it does in extensor digitorum longus and epitrochlearis muscles, 2) the ability of AICAR to increase fatty acid oxidation and diminish glucose oxidation and malonyl-CoA concentration is dependent on the nutritional status of the rat, and 3) the ability of AICAR to diminish assayable ACC activity is independent of nutritional state. (+info)
Regulation of muscle GLUT-4 transcription by AMP-activated protein kinase.
(31/623)
Skeletal muscle GLUT-4 transcription in response to treatment with 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside (AICAR), a known activator of AMP-activated protein kinase (AMPK), was studied in rats and mice. The increase in GLUT-4 mRNA levels in response to a single subcutaneous injection of AICAR, peaked at 13 h in white and red quadriceps muscles but not in the soleus muscle. The mRNA level of chloramphenicol acyltransferase reporter gene which is driven by 1,154 or 895 bp of the human GLUT-4 proximal promoter was increased in AICAR-treated transgenic mice, demonstrating the transcriptional upregulation of the GLUT-4 gene by AICAR. However, this induction of transcription was not apparent with 730 bp of the promoter. In addition, nuclear extracts from AICAR-treated mice bound to the consensus sequence of myocyte enhancer factor-2 (from -473 to -464) to a greater extent than from saline-injected mice. Thus AMP-activated protein kinase activation by AICAR increases GLUT-4 transcription by a mechanism that requires response elements within 895 bp of human GLUT-4 proximal promoter and that may be cooperatively mediated by myocyte enhancer factor-2. (+info)
Stimulation of glucose transport by AMP-activated protein kinase via activation of p38 mitogen-activated protein kinase.
(32/623)
Activation of AMP-activated protein kinase (AMPK) has been recently demonstrated to be associated with 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR)-stimulated glucose transport mediated by both GLUT1 and GLUT4 transporters. However, signaling events upstream and downstream of AMPK are unknown. Here we report that 1) p38 mitogen-activated protein kinase (MAPK) and mitogen-activated protein kinase kinase 3 (MKK3) were activated by AICAR in Clone 9 cells, which express only the GLUT1 transporters, and 2) activation of p38 was required for AICAR-stimulated glucose transport since treatment of the cells with p38 inhibitor SB203580 or overexpression of dominant negative p38 mutant inhibited glucose transport. Moreover, we found that overexpression of the constitutively active form of AMPK mutant also resulted in a significant activation of p38, and inhibition of p38 activity by SB203580 did not affect AICAR-stimulated activation of AMPK. These findings demonstrate that AICAR-stimulated activation of p38 is indeed mediated by AMPK, and the p38 MAPK cascade is downstream of AMPK in the signaling pathway of AICAR-stimulated glucose transport in Clone 9 cells. (+info)