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Robert A Harris - One of the best experts on this subject based on the ideXlab platform.
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Pyruvate Dehydrogenase Kinase-Mediated Glycolytic Metabolic Shift in the Dorsal Root Ganglion Drives Painful Diabetic Neuropathy
The Journal of biological chemistry, 2016Co-Authors: Habibur Rahman, Robert A Harris, Mithilesh Kumar Jha, Dong Ho Park, Hyun Kook, Maan-gee Lee, Jong-heon Kim, Youngpyo Nam, In-kyu LeeAbstract:The dorsal root ganglion (DRG) is a highly vulnerable site in diabetic neuropathy. Under diabetic conditions, the DRG is subjected to tissue ischemia or lower ambient oxygen tension that leads to aberrant metabolic functions. Metabolic dysfunctions have been documented to play a crucial role in the pathogenesis of diverse pain hypersensitivities. However, the contribution of diabetes-induced metabolic dysfunctions in the DRG to the pathogenesis of painful diabetic neuropathy remains ill-explored. In this study, we report that pyruvate dehydrogenase kinases (PDK2 and PDK4), key regulatory enzymes in glucose metabolism, mediate glycolytic metabolic shift in the DRG leading to painful diabetic neuropathy. Streptozotocin-induced diabetes substantially enhanced the expression and activity of the PDKs in the DRG, and the genetic ablation of PDK2 and Pdk4 attenuated the hyperglycemia-induced pain hypersensitivity. Mechanistically, PDK2/4 deficiency inhibited the diabetes-induced lactate surge, expression of pain-related ion channels, activation of satellite glial cells, and infiltration of macrophages in the DRG, in addition to reducing central sensitization and neuroinflammation hallmarks in the spinal cord, which probably accounts for the attenuated pain hypersensitivity. PDK2/4-deficient mice were partly resistant to the diabetes-induced loss of peripheral nerve structure and function. Furthermore, in the experiments using DRG neuron cultures, lactic acid treatment enhanced the expression of the ion channels and compromised cell viability. Finally, the pharmacological inhibition of DRG PDKs or lactic acid production substantially attenuated diabetes-induced pain hypersensitivity. Taken together, PDK2/4 induction and the subsequent lactate surge induce the metabolic shift in the diabetic DRG, thereby contributing to the pathogenesis of painful diabetic neuropathy.
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Metabolic Connection of Inflammatory Pain: Pivotal Role of a Pyruvate Dehydrogenase Kinase-Pyruvate Dehydrogenase-Lactic Acid Axis.
The Journal of neuroscience : the official journal of the Society for Neuroscience, 2015Co-Authors: Mithilesh Kumar Jha, Nam Ho Jeoung, Robert A Harris, Dong Ho Park, Hyun Kook, In-kyu Lee, Gyun Jee Song, Maan-gee Lee, Kyoungho SukAbstract:Pyruvate dehydrogenase kinases (PDK1–4) are mitochondrial metabolic regulators that serve as decision makers via modulation of pyruvate dehydrogenase (PDH) activity to convert pyruvate either aerobically to acetyl-CoA or anaerobically to lactate. Metabolic dysregulation and inflammatory processes are two sides of the same coin in several pathophysiological conditions. The lactic acid surge associated with the metabolic shift has been implicated in diverse painful states. In this study, we investigated the role of PDK-PDH-lactic acid axis in the pathogenesis of chronic inflammatory pain. Deficiency of PDK2 and/or Pdk4 in mice attenuated complete Freund9s adjuvant (CFA)-induced pain hypersensitivities. Likewise, PDK2/4 deficiency attenuated the localized lactic acid surge along with hallmarks of peripheral and central inflammation following intraplantar administration of CFA. In vitro studies supported the role of PDK2/4 as promoters of classical proinflammatory activation of macrophages. Moreover, the pharmacological inhibition of PDKs or lactic acid production diminished CFA-induced inflammation and pain hypersensitivities. Thus, a PDK-PDH-lactic acid axis seems to mediate inflammation-driven chronic pain, establishing a connection between metabolism and inflammatory pain. SIGNIFICANCE STATEMENT The mitochondrial pyruvate dehydrogenase (PDH) kinases (PDKs) and their substrate PDH orchestrate the conversion of pyruvate either aerobically to acetyl-CoA or anaerobically to lactate. Lactate, the predominant end product of glycolysis, has recently been identified as a signaling molecule for neuron-glia interactions and neuronal plasticity. Pathological metabolic shift and subsequent lactic acid production are thought to play an important role in diverse painful states; however, their contribution to inflammation-driven pain is still to be comprehended. Here, we report that the PDK-PDH-lactic acid axis constitutes a key component of inflammatory pain pathogenesis. Our findings establish an unanticipated link between metabolism and inflammatory pain. This study unlocks a previously ill-explored research avenue for the metabolic control of inflammatory pain pathogenesis.
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Genetic inactivation of pyruvate dehydrogenase kinases improves hepatic insulin resistance induced diabetes.
PloS one, 2013Co-Authors: Rongya Tao, Robert A Harris, Xiwen Xiong, Morris F. White, Xiaocheng C. DongAbstract:Pyruvate dehydrogenase kinases (PDK1-4) play a critical role in the inhibition of the mitochondrial pyruvate dehydrogenase complex especially when blood glucose levels are low and pyruvate can be conserved for gluconeogenesis. Under diabetic conditions, the Pdk genes, particularly Pdk4, are often induced, and the elevation of the Pdk4 gene expression has been implicated in the increased gluconeogenesis in the liver and the decreased glucose utilization in the peripheral tissues. However, there is no direct evidence yet to show to what extent that the dysregulation of hepatic Pdk genes attributes to hyperglycemia and insulin resistance in vivo. To address this question, we crossed PDK2 or Pdk4 null mice with a diabetic model that is deficient in hepatic insulin receptor substrates 1 and 2 (Irs1/2). Metabolic analyses reveal that deletion of the Pdk4 gene had better improvement in hyperglycemia and glucose tolerance than knockout of the PDK2 gene whereas the PDK2 gene deletion showed better insulin tolerance as compared to the Pdk4 gene inactivation on the Irs1/2 knockout genetic background. To examine the specific hepatic effects of Pdks on diabetes, we also knocked down the PDK2 or Pdk4 gene using specific shRNAs. The data also indicate that the Pdk4 gene knockdown led to better glucose tolerance than the PDK2 gene knockdown. In conclusion, our data suggest that hepatic Pdk4 may be critically involved in the pathogenesis of diabetes.
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PDH activation during in vitro muscle contractions in PDH kinase 2 knockout mice: effect of PDH kinase 1 compensation
American Journal of Physiology-regulatory Integrative and Comparative Physiology, 2011Co-Authors: Emily C.e. Dunford, Nam Ho Jeoung, Robert A Harris, Eric A.f. Herbst, William Gittings, J. Greig Inglis, Rene Vandenboom, Paul J. Leblanc, Sandra J. PetersAbstract:Pyruvate dehydrogenase (PDH) plays an important role in regulating carbohydrate oxidation in skeletal muscle. PDH is deactivated by a set of PDH kinases (PDK1, PDK2, PDK3, PDK4), with PDK2 and PDK4...
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PDH activation during in vitro muscle contractions in PDH kinase 2 knockout mice: effect of PDH kinase 1 compensation.
American journal of physiology. Regulatory integrative and comparative physiology, 2011Co-Authors: Emily C.e. Dunford, Nam Ho Jeoung, Robert A Harris, Eric A.f. Herbst, William Gittings, J. Greig Inglis, Rene Vandenboom, Paul J. Leblanc, Sandra J. PetersAbstract:Pyruvate dehydrogenase (PDH) plays an important role in regulating carbohydrate oxidation in skeletal muscle. PDH is deactivated by a set of PDH kinases (PDK1, PDK2, PDK3, PDK4), with PDK2 and PDK4 being the most predominant isoforms in skeletal muscle. Although PDK2 is the most abundant isoform, few studies have examined its physiological role. The role of PDK2 on PDH activation (PDHa) at rest and during muscle stimulation at 10 and 40 Hz (eliciting low- and moderate-intensity muscle contractions, respectively) in isolated extensor digitorum longus muscles was studied in PDK2 knockout (PDK2KO) and wild-type (WT) mice (n = 5 per group). PDHa activity was unexpectedly 35 and 77% lower in PDK2KO than WT muscle (P = 0.043), while total PDK activity was nearly fourfold lower in PDK2KO muscle (P = 0.006). During 40-Hz contractions, initial force was lower in PDK2KO than WT muscle (P < 0.001) but fatigued similarly to ∼75% of initial force by 3 min. There were no differences in initial force or rate of fatigue during 10-Hz contractions. PDK1 compensated for the lack of PDK2 and was 1.8-fold higher in PDK2KO than WT muscle (P = 0.019). This likely contributed to ensuring that resting PDHa activity was similar between the groups and accounts for the lower PDH activation during muscle contraction, as PDK1 is a very potent inhibitor of the PDH complex. Increased PDK1 expression appears to be regulated by hypoxia inducible factor-1α, which was 3.5-fold higher in PDK2KO muscle. It is clear that PDK2 activity is essential, even at rest, in regulation of carbohydrate oxidation and production of reducing equivalents for the electron transport chain. In addition, these results underscore the importance of the overall kinetics of the PDK isoform population, rather than total PDK activity, in determining transformation of the PDH complex and PDHa activity during muscle contraction.
Mary C Sugden - One of the best experts on this subject based on the ideXlab platform.
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Mechanisms underlying regulation of the expression and activities of the mammalian pyruvate dehydrogenase kinases
Archives of Physiology and Biochemistry, 2006Co-Authors: Mary C Sugden, Mark J HolnessAbstract:AbstractThe mechanisms that control mammalian pyruvate dehydrogenase complex (PDC) activity include its phosphorylation (inactivation) by a family of pyruvate dehydrogenase kinases (PDKs 1 – 4). Here we review new developments in the regulation of the activities and expression of the PDKs, in particular PDK2 and PDK4, in relation to glucose and lipid homeostasis. This review describes recent advances relating to the acute and long-term modes of regulation of the PDKs, with particular emphasis on the regulatory roles of nuclear receptors including peroxisome proliferator-activated receptor (PPAR) α and Liver X receptor (LXR), PPAR γ coactivator α (PGC-1α) and insulin, and the impact of changes in PDK activity and expression in glucose and lipid homeostasis. Since PDK4 may assist in lipid clearance when there is an imbalance between lipid delivery and oxidation, it may represent an attractive target for interventions aimed at rectifying abnormal lipid as well as glucose homeostasis in disease states.
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investigation of potential mechanisms regulating protein expression of hepatic pyruvate dehydrogenase kinase isoforms 2 and 4 by fatty acids and thyroid hormone
Biochemical Journal, 2003Co-Authors: Mark J Holness, Karen Bulmer, Nicholas D Smith, Mary C SugdenAbstract:: Liver contains two pyruvate dehydrogenase kinases (PDKs), namely PDK2 and PDK4, which regulate glucose oxidation through inhibitory phosphorylation of the pyruvate dehydrogenase complex (PDC). Starvation increases hepatic PDK2 and PDK4 protein expression, the latter occurring, in part, via a mechanism involving peroxisome proliferator-activated receptor-alpha (PPARalpha). High-fat feeding and hyperthyroidism, which increase circulating lipid supply, enhance hepatic PDK2 protein expression, but these increases are insufficient to account for observed increases in hepatic PDK activity. Enhanced expression of PDK4, but not PDK2, occurs in part via a mechanism involving PPAR-alpha. Heterodimerization partners for retinoid X receptors (RXRs) include PPARalpha and thyroid-hormone receptors (TRs). We therefore investigated the responses of hepatic PDK protein expression to high-fat feeding and hyperthyroidism in relation to hepatic lipid delivery and disposal. High-fat feeding increased hepatic PDK2, but not PDK4, protein expression whereas hyperthyroidism increased both hepatic PDK2 and PDK4 protein expression. Both manipulations decreased the sensitivity of hepatic carnitine palmitoyltransferase I (CPT I) to suppression by malonyl-CoA, but only hyperthyrodism elevated plasma fatty acid and ketone-body concentrations and CPT I maximal activity. Administration of the selective PPAR-alpha activator WY14,643 significantly increased PDK4 protein to a similar extent in both control and high-fat-fed rats, but WY14,643 treatment and hyperthyroidism did not have additive effects on hepatic PDK4 protein expression. PPARalpha activation did not influence hepatic PDK2 protein expression in euthyroid rats, suggesting that up-regulation of PDK2 by hyperthyroidism does not involve PPARalpha, but attenuated the effect of hyperthyroidism to increase hepatic PDK2 expression. The results indicate that hepatic PDK4 up-regulation can be achieved by heterodimerization of either PPARalpha or TR with the RXR receptor and that effects of PPARalpha activation on hepatic PDK2 and PDK4 expression favour a switch towards preferential expression of PDK4.
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Selective Modification of Pyruvate Dehydrogenase Kinase Isoform Expression in Rat Pancreatic Islets Elicited by Starvation and Activation of Peroxisome Proliferator–Activated Receptor-α: Implications for Glucose-Stimulated Insulin Secretion
Diabetes, 2001Co-Authors: Mary C Sugden, Karen Bulmer, Daniel Augustine, Mark J HolnessAbstract:The pyruvate dehydrogenase complex (PDC) has a pivotal role in islet metabolism. The pyruvate dehydrogenase kinases (PDK1–4) regulate glucose oxidation through inhibitory phosphorylation of PDC. Starvation increases islet PDK activity (AmJ Physiol Endocrinol Metab 270:E988–E994, 1996). In this study, using antibodies against PDK1, PDK2, and PDK4 (no sufficiently specific antibodies are as yet available for PDK3), we identified the PDK isoform profile of the pancreatic islet and delineated the effects of starvation (48 h) on protein expression of individual PDK isoforms. Rat islets were demonstrated to contain all three PDK isoforms, PDK1, PDK2, and PDK4. Using immunoblot analysis with antibodies raised against the individual recombinant PDK isoforms, we demonstrated increased islet protein expression of PDK4 in response to starvation (2.3-fold; P
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targeted upregulation of pyruvate dehydrogenase kinase pdk 4 in slow twitch skeletal muscle underlies the stable modification of the regulatory characteristics of pdk induced by high fat feeding
Diabetes, 2000Co-Authors: Mark J Holness, A Kraus, Robert A Harris, Mary C SugdenAbstract:In using Western blot analysis with antibodies raised against recombinant pyruvate dehydrogenase kinase (PDK) isoforms PDK2 and PDK4, this study demonstrates selective PDK isoform switching in specific skeletal muscle types in response to high-fat feeding that is associated with altered regulation of PDK activity by pyruvate. The administration of a diet high in saturated fats led to stable (approximately 2-fold) increases in PDK activities in both a typical slow-twitch (soleus [SOL]) muscle and a typical fast-twitch (anterior tibialis [AT]) muscle. Western blot analysis revealed that high-fat feeding significantly increased (approximately 2-fold; P
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Targeted upregulation of pyruvate dehydrogenase kinase (PDK)-4 in slow-twitch skeletal muscle underlies the stable modification of the regulatory characteristics of PDK induced by high-fat feeding.
Diabetes, 2000Co-Authors: Mark J Holness, A Kraus, Robert A Harris, Mary C SugdenAbstract:In using Western blot analysis with antibodies raised against recombinant pyruvate dehydrogenase kinase (PDK) isoforms PDK2 and PDK4, this study demonstrates selective PDK isoform switching in specific skeletal muscle types in response to high-fat feeding that is associated with altered regulation of PDK activity by pyruvate. The administration of a diet high in saturated fats led to stable (approximately 2-fold) increases in PDK activities in both a typical slow-twitch (soleus [SOL]) muscle and a typical fast-twitch (anterior tibialis [AT]) muscle. Western blot analysis revealed that high-fat feeding significantly increased (approximately 2-fold; P < 0.001) PDK4 protein expression in SOL, with a modest (1.3-fold) increase in PDK2 protein expression. The relative increase in PDK4 protein expression in SOL was associated with a 7.6-fold increase in the pyruvate concentration that was required to elicit a 50% active pyruvate dehydrogenase complex, which indicates a marked decrease in the sensitivity of PDK to inhibition by pyruvate. In AT muscle, high-fat feeding elicited comparable (1.5- to 1.7-fold) increases (P < 0.05) in PDK4 and PDK2 protein expression. Loss of sensitivity of PDK to inhibition by pyruvate was less marked. The data suggest that a positive correlation exists between increases in PDK4 expression and the propensity with which muscles use lipid-derived fuels as respiratory substrates rather than with the degree of insulin resistance induced in skeletal muscles by high-fat feeding. In conclusion, high-fat feeding leads to selective upregulation of PDK4 expression in slow-twitch muscle in response to high-fat feeding in vivo, which is associated with a pronounced loss of sensitivity of PDK activity to acute inhibition by pyruvate. Thus, increased PDK4 expression may underlie the stable modification of the regulatory characteristics of PDK observed in slow-twitch muscle in response to high-fat feeding.
Eungbin Kim - One of the best experts on this subject based on the ideXlab platform.
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Heat Shock-Induced Physical Changes of Megaplasmids in Rhodococcus sp. Strain DK17
2011Co-Authors: Kyungsun Kim, Dockyu Kim, Hae-youn Park, Junghee Sung, Eungbin KimAbstract:Rhodococcus sp. strain DK17 possesses three megaplasmids (380 kb pDK1, 330 kb PDK2, and 750 kb pDK3). The alkylbenzene-degrading genes (akbABCDEF) are present on PDK2 while the phthalate operons which are duplicated are present on both PDK2 (ophA’B’C’R’) and pDK3 (ophABCR). DK17 with an optimal temperature of 30°C showed no growth at 37°C. When transferred to 30°C, however, the 37°C culture began to grow immediately, indicating that 37°C is not lethal but stressful for DK17 growth. In addition, when exposed to 37°C even for a short time, a part of DK17 cells lost the ability to degrade o-xylene (a model compound of alkylbenzenes). When two hundred colonies were randomly selected for colony PCR for PDK2-specific akbC, ophC’, or pDK3-specific ophC, a total of 29 colonies were found to have lost at least one of the three genes. PFGE analysis clearly showed that all the mutants have different megaplasmid profiles from that of DK17 wild type, which are divided into five different cases: Type I (10 mutants, PDK2 loss and acquisition of a new ~700 kb plasmid), Type II (9 mutants, PDK2 loss), Type III (8 mutants, pDK3 loss and acquisition of a new ~400 kb plasmid), Type IV (1 mutant, pDK3 loss), and Type V (1 mutant, PDK2 and pDK3 loss and acquisition of the ~400 kb and ~700 kb plasmids). The above results showing that growth temperature changes can induce physical changes in bacterial genomes suggest that environmental changes in habitats including temperature fluctuations affect significantly the evolution of bacteria.Keywords: Rhodococcus, heat-shock, megaplasmid, PFGE
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Requirement of duplicated operons for maximal metabolism of phthalate by Rhodococcus sp. strain DK17.
Biochemical and biophysical research communications, 2007Co-Authors: Ki Young Choi, Dockyu Kim, Jong-chan Chae, Gerben J. Zylstra, Eungbin KimAbstract:Abstract The operons encoding the transformation of phthalate to protocatechuate are duplicated and present on two different megaplasmids [PDK2 (330 kb) and pDK3 (750 kb)] in Rhodococcus sp. strain DK17. RT-PCR experiments using gene-specific primers showed that both the PDK2- and the pDK3-encoded dihydroxyphthalate decarboxylase genes are simultaneously expressed during growth on phthalate. The doubling time of the PDK2-cured mutant strain DK176 in minimal liquid medium with 5 mM phthalate is 52.5% of that of the wild-type strain DK17. The data indicate that both copies of the phthalate operon are equally functional in DK17, and gene dosage is the main reason for slower growth of DK176 on phthalate.
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Molecular and biochemical analysis of phthalate and terephthalate degradation by Rhodococcus sp. strain DK17.
FEMS microbiology letters, 2005Co-Authors: Ki Young Choi, Dockyu Kim, Woo Jun Sul, Jong-chan Chae, Gerben J. Zylstra, Young Min Kim, Eungbin KimAbstract:Alkylbenzene-degrading Rhodococcus sp. strain DK17 is able to utilize phthalate and terephthalate as growth substrates. The genes encoding the transformation of phthalate and terephthalate to protocatechuate are organized as two separate operons, located 6.7 kb away from each other. Interestingly, both the phthalate and terephthalate operons are induced in response to terephthalate while expression of the terephthalate genes is undetectable in phthalate-grown cells. In addition to two known plasmids (380-kb pDK1 and 330-kb PDK2), a third megaplasmid (750-kb pDK3) was newly identified in DK17. The phthalate and terephthalate operons are duplicated and are present on both PDK2 and pDK3. RT-PCR experiments, coupled with sequence analysis, suggest that phthalate and terephthalate degradation in DK17 proceeds through oxygenation at carbons 3 and 4 and at carbons 1 and 2 to form 3,4-dihydro-3,4-dihydroxyphthalate and 1,2-dihydro-1,2-dihydroxyterephthalate, respectively. The 3,4-dihydroxyphthalate pathway was further corroborated through colorometric tests. Apparently, the two dihydrodiol metabolites are subsequently dehydrogenated and decarboxylated to form protocatechuate, which is further degraded by a protocatechuate 3,4-dioxygenase as confirmed by a ring-cleavage enzyme assay.
Mark J Holness - One of the best experts on this subject based on the ideXlab platform.
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Mechanisms underlying regulation of the expression and activities of the mammalian pyruvate dehydrogenase kinases
Archives of Physiology and Biochemistry, 2006Co-Authors: Mary C Sugden, Mark J HolnessAbstract:AbstractThe mechanisms that control mammalian pyruvate dehydrogenase complex (PDC) activity include its phosphorylation (inactivation) by a family of pyruvate dehydrogenase kinases (PDKs 1 – 4). Here we review new developments in the regulation of the activities and expression of the PDKs, in particular PDK2 and PDK4, in relation to glucose and lipid homeostasis. This review describes recent advances relating to the acute and long-term modes of regulation of the PDKs, with particular emphasis on the regulatory roles of nuclear receptors including peroxisome proliferator-activated receptor (PPAR) α and Liver X receptor (LXR), PPAR γ coactivator α (PGC-1α) and insulin, and the impact of changes in PDK activity and expression in glucose and lipid homeostasis. Since PDK4 may assist in lipid clearance when there is an imbalance between lipid delivery and oxidation, it may represent an attractive target for interventions aimed at rectifying abnormal lipid as well as glucose homeostasis in disease states.
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investigation of potential mechanisms regulating protein expression of hepatic pyruvate dehydrogenase kinase isoforms 2 and 4 by fatty acids and thyroid hormone
Biochemical Journal, 2003Co-Authors: Mark J Holness, Karen Bulmer, Nicholas D Smith, Mary C SugdenAbstract:: Liver contains two pyruvate dehydrogenase kinases (PDKs), namely PDK2 and PDK4, which regulate glucose oxidation through inhibitory phosphorylation of the pyruvate dehydrogenase complex (PDC). Starvation increases hepatic PDK2 and PDK4 protein expression, the latter occurring, in part, via a mechanism involving peroxisome proliferator-activated receptor-alpha (PPARalpha). High-fat feeding and hyperthyroidism, which increase circulating lipid supply, enhance hepatic PDK2 protein expression, but these increases are insufficient to account for observed increases in hepatic PDK activity. Enhanced expression of PDK4, but not PDK2, occurs in part via a mechanism involving PPAR-alpha. Heterodimerization partners for retinoid X receptors (RXRs) include PPARalpha and thyroid-hormone receptors (TRs). We therefore investigated the responses of hepatic PDK protein expression to high-fat feeding and hyperthyroidism in relation to hepatic lipid delivery and disposal. High-fat feeding increased hepatic PDK2, but not PDK4, protein expression whereas hyperthyroidism increased both hepatic PDK2 and PDK4 protein expression. Both manipulations decreased the sensitivity of hepatic carnitine palmitoyltransferase I (CPT I) to suppression by malonyl-CoA, but only hyperthyrodism elevated plasma fatty acid and ketone-body concentrations and CPT I maximal activity. Administration of the selective PPAR-alpha activator WY14,643 significantly increased PDK4 protein to a similar extent in both control and high-fat-fed rats, but WY14,643 treatment and hyperthyroidism did not have additive effects on hepatic PDK4 protein expression. PPARalpha activation did not influence hepatic PDK2 protein expression in euthyroid rats, suggesting that up-regulation of PDK2 by hyperthyroidism does not involve PPARalpha, but attenuated the effect of hyperthyroidism to increase hepatic PDK2 expression. The results indicate that hepatic PDK4 up-regulation can be achieved by heterodimerization of either PPARalpha or TR with the RXR receptor and that effects of PPARalpha activation on hepatic PDK2 and PDK4 expression favour a switch towards preferential expression of PDK4.
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Selective Modification of Pyruvate Dehydrogenase Kinase Isoform Expression in Rat Pancreatic Islets Elicited by Starvation and Activation of Peroxisome Proliferator–Activated Receptor-α: Implications for Glucose-Stimulated Insulin Secretion
Diabetes, 2001Co-Authors: Mary C Sugden, Karen Bulmer, Daniel Augustine, Mark J HolnessAbstract:The pyruvate dehydrogenase complex (PDC) has a pivotal role in islet metabolism. The pyruvate dehydrogenase kinases (PDK1–4) regulate glucose oxidation through inhibitory phosphorylation of PDC. Starvation increases islet PDK activity (AmJ Physiol Endocrinol Metab 270:E988–E994, 1996). In this study, using antibodies against PDK1, PDK2, and PDK4 (no sufficiently specific antibodies are as yet available for PDK3), we identified the PDK isoform profile of the pancreatic islet and delineated the effects of starvation (48 h) on protein expression of individual PDK isoforms. Rat islets were demonstrated to contain all three PDK isoforms, PDK1, PDK2, and PDK4. Using immunoblot analysis with antibodies raised against the individual recombinant PDK isoforms, we demonstrated increased islet protein expression of PDK4 in response to starvation (2.3-fold; P
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targeted upregulation of pyruvate dehydrogenase kinase pdk 4 in slow twitch skeletal muscle underlies the stable modification of the regulatory characteristics of pdk induced by high fat feeding
Diabetes, 2000Co-Authors: Mark J Holness, A Kraus, Robert A Harris, Mary C SugdenAbstract:In using Western blot analysis with antibodies raised against recombinant pyruvate dehydrogenase kinase (PDK) isoforms PDK2 and PDK4, this study demonstrates selective PDK isoform switching in specific skeletal muscle types in response to high-fat feeding that is associated with altered regulation of PDK activity by pyruvate. The administration of a diet high in saturated fats led to stable (approximately 2-fold) increases in PDK activities in both a typical slow-twitch (soleus [SOL]) muscle and a typical fast-twitch (anterior tibialis [AT]) muscle. Western blot analysis revealed that high-fat feeding significantly increased (approximately 2-fold; P
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Targeted upregulation of pyruvate dehydrogenase kinase (PDK)-4 in slow-twitch skeletal muscle underlies the stable modification of the regulatory characteristics of PDK induced by high-fat feeding.
Diabetes, 2000Co-Authors: Mark J Holness, A Kraus, Robert A Harris, Mary C SugdenAbstract:In using Western blot analysis with antibodies raised against recombinant pyruvate dehydrogenase kinase (PDK) isoforms PDK2 and PDK4, this study demonstrates selective PDK isoform switching in specific skeletal muscle types in response to high-fat feeding that is associated with altered regulation of PDK activity by pyruvate. The administration of a diet high in saturated fats led to stable (approximately 2-fold) increases in PDK activities in both a typical slow-twitch (soleus [SOL]) muscle and a typical fast-twitch (anterior tibialis [AT]) muscle. Western blot analysis revealed that high-fat feeding significantly increased (approximately 2-fold; P < 0.001) PDK4 protein expression in SOL, with a modest (1.3-fold) increase in PDK2 protein expression. The relative increase in PDK4 protein expression in SOL was associated with a 7.6-fold increase in the pyruvate concentration that was required to elicit a 50% active pyruvate dehydrogenase complex, which indicates a marked decrease in the sensitivity of PDK to inhibition by pyruvate. In AT muscle, high-fat feeding elicited comparable (1.5- to 1.7-fold) increases (P < 0.05) in PDK4 and PDK2 protein expression. Loss of sensitivity of PDK to inhibition by pyruvate was less marked. The data suggest that a positive correlation exists between increases in PDK4 expression and the propensity with which muscles use lipid-derived fuels as respiratory substrates rather than with the degree of insulin resistance induced in skeletal muscles by high-fat feeding. In conclusion, high-fat feeding leads to selective upregulation of PDK4 expression in slow-twitch muscle in response to high-fat feeding in vivo, which is associated with a pronounced loss of sensitivity of PDK activity to acute inhibition by pyruvate. Thus, increased PDK4 expression may underlie the stable modification of the regulatory characteristics of PDK observed in slow-twitch muscle in response to high-fat feeding.
Ki Young Choi - One of the best experts on this subject based on the ideXlab platform.
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Requirement of duplicated operons for maximal metabolism of phthalate by Rhodococcus sp. strain DK17.
Biochemical and biophysical research communications, 2007Co-Authors: Ki Young Choi, Dockyu Kim, Jong-chan Chae, Gerben J. Zylstra, Eungbin KimAbstract:Abstract The operons encoding the transformation of phthalate to protocatechuate are duplicated and present on two different megaplasmids [PDK2 (330 kb) and pDK3 (750 kb)] in Rhodococcus sp. strain DK17. RT-PCR experiments using gene-specific primers showed that both the PDK2- and the pDK3-encoded dihydroxyphthalate decarboxylase genes are simultaneously expressed during growth on phthalate. The doubling time of the PDK2-cured mutant strain DK176 in minimal liquid medium with 5 mM phthalate is 52.5% of that of the wild-type strain DK17. The data indicate that both copies of the phthalate operon are equally functional in DK17, and gene dosage is the main reason for slower growth of DK176 on phthalate.
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Molecular and biochemical analysis of phthalate and terephthalate degradation by Rhodococcus sp. strain DK17.
FEMS microbiology letters, 2005Co-Authors: Ki Young Choi, Dockyu Kim, Woo Jun Sul, Jong-chan Chae, Gerben J. Zylstra, Young Min Kim, Eungbin KimAbstract:Alkylbenzene-degrading Rhodococcus sp. strain DK17 is able to utilize phthalate and terephthalate as growth substrates. The genes encoding the transformation of phthalate and terephthalate to protocatechuate are organized as two separate operons, located 6.7 kb away from each other. Interestingly, both the phthalate and terephthalate operons are induced in response to terephthalate while expression of the terephthalate genes is undetectable in phthalate-grown cells. In addition to two known plasmids (380-kb pDK1 and 330-kb PDK2), a third megaplasmid (750-kb pDK3) was newly identified in DK17. The phthalate and terephthalate operons are duplicated and are present on both PDK2 and pDK3. RT-PCR experiments, coupled with sequence analysis, suggest that phthalate and terephthalate degradation in DK17 proceeds through oxygenation at carbons 3 and 4 and at carbons 1 and 2 to form 3,4-dihydro-3,4-dihydroxyphthalate and 1,2-dihydro-1,2-dihydroxyterephthalate, respectively. The 3,4-dihydroxyphthalate pathway was further corroborated through colorometric tests. Apparently, the two dihydrodiol metabolites are subsequently dehydrogenated and decarboxylated to form protocatechuate, which is further degraded by a protocatechuate 3,4-dioxygenase as confirmed by a ring-cleavage enzyme assay.