Regulation of muscle creatine kinase by phosphorylation in normal and diabetic hearts

Lin, Guorong; Liu, Y.; MacLeod, Kathleen M.

doi:10.1007/s00018-008-8575-3

Cited by 25 publications

(24 citation statements)

References 33 publications

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“…All major PKC isoforms are also detected in mitochondria isolated by Percoll purification from normal hearts [31]. We and others have reported that levels of PKCb 2 and PKCe are increased in total cellular protein from diabetic hearts [11,12,19]. However, whether this is associated with changes in the levels of these isoforms in mitochondria from diabetic hearts is not known.…”

Section: Resultsmentioning

confidence: 96%

“…1A), an 80-kDa protein with increased phosphorylation and a 40-kDa protein with reduced phosphorylation, in hearts from diabetic rats. The 40-kDa protein was identified as M-CK, and further characterization was recently reported [19]. To identify the 80-kDa protein, the corresponding band was cut from the gel, subjected to in-gel digestion and the resulting peptides were analyzed by MS.…”

Section: Resultsmentioning

confidence: 99%

“…The experiments were carried out as described in our recent study [19]. The candidate proteins were identified by Western blotting with antiphospho-(Ser) PKC substrate antibody.…”

Section: Western Blotting Analysis and Immunoprecipitationmentioning

confidence: 99%

“…Using an antibody that selectively recognizes a phosphoserine PKC substrate motif, we recently found reciprocal changes in PKC-mediated phosphorylation of two proteins in diabetic hearts. The protein with decreased phosphorylation was identified, by mass spectrometry (MS), as muscle creatine kinase (M-CK) [19]. The reduced PKCmediated phosphorylation of Ser128 in M-CK causes a reduction in the forward reaction that transfers highenergy phosphate from ATP to generate phosphocreatine, and an increase in the reverse reaction that shuttles high-energy phosphate from phosphocreatine to ATP.…”

Section: Introductionmentioning

confidence: 99%

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Regulation of mitochondrial aconitase by phosphorylation in diabetic rat heart

Lin

Brownsey

MacLeod

2009

Cell. Mol. Life Sci.

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Mitochondrial dysfunction and protein kinase C (PKC) activation are consistently found in diabetic cardiomyopathy but their relationship remains unclear. This study identified mitochondrial aconitase as a downstream target of PKC activation using immunoblotting and mass spectrometry, and then characterized phosphorylation-induced changes in its activity in hearts from type 1 diabetic rats. PKCbeta(2) co-immunoprecipitated with phosphorylated aconitase from mitochondria isolated from diabetic hearts. Augmented phosphorylation of mitochondrial aconitase in diabetic hearts was found to be associated with an increase in its reverse activity (isocitrate to aconitate), while the rate of the forward activity was unchanged. Similar results were obtained on phosphorylation of mitochondrial aconitase by PKCbeta(2) in vitro. These results demonstrate the regulation of mitochondrial aconitase activity by PKC-dependent phosphorylation. This may influence the activity of the tricarboxylic acid cycle, and contribute to impaired mitochondrial function and energy metabolism in diabetic hearts.

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Section: Resultsmentioning

confidence: 96%

Section: Resultsmentioning

confidence: 99%

“…The experiments were carried out as described in our recent study [19]. The candidate proteins were identified by Western blotting with antiphospho-(Ser) PKC substrate antibody.…”

Section: Western Blotting Analysis and Immunoprecipitationmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Regulation of mitochondrial aconitase by phosphorylation in diabetic rat heart

Lin

Brownsey

MacLeod

2009

Cell. Mol. Life Sci.

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“…A Putative PKC Phosphorylation Site on sMiCK and CKM Is Required for the Regulation of NCX1 Activity-It has been shown that CKM is phosphorylated by PKC, possibly at Ser-128 (32). To examine the role of this serine residue in CK mediated recovery of the decreased NCX1 activity, plasmids encoding the Ser-128 equivalent mutants of sMiCK and CKM, namely sMiCK-S123A and CKM-S128A, were constructed and co-transfected with NCX1 into HEK293T cells.…”

Section: Regulation Of Namentioning

confidence: 99%

Regulation of Sodium-Calcium Exchanger Activity by Creatine Kinase under Energy-compromised Conditions

Yang¹,

Fann²,

Chang³

et al. 2010

Journal of Biological Chemistry

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Na؉ /Ca 2؉ exchanger (NCX) is one of the major mechanisms for removing Ca 2؉ from the cytosol especially in cardiac myocytes and neurons, where their physiological activities are triggered by an influx of Ca 2؉ . NCX contains a large intracellular loop (NCXIL) that is responsible for regulating NCX activity. Recent evidence has shown that proteins, including kinases and phosphatases, associate with NCX1IL to form a NCX1 macromolecular complex. To search for the molecules that interact with NCX1IL and regulate NCX1 activity, we used the yeast twohybrid method to screen a human heart cDNA library and found that the C-terminal region of sarcomeric mitochondrial creatine kinase (sMiCK) interacted with NCX1IL. Moreover, both sMiCK and the muscle-type creatine kinase (CKM) coimmunoprecipitated with NCX1 using lysates of cardiacmyocytes and HEK293T cells that transiently expressed NCX1 and various creatine kinases. Both sMiCK and CKM were able to produce a recovery in the decreased NCX1 activity that was lost under energy-compromised conditions. This regulation is mediated through a putative PKC phosphorylation site of sMiCK and CKM. The autophosphorylation and the catalytic activity of sMiCK and CKM are not required for their regulation of NCX1 activity. Our results suggest a novel mechanism for the regulation of NCX1 activity.

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Regulation of Sodium-Calcium Exchanger Activity by Creatine Kinase

Yang¹,

Kao²

2012

Advances in Experimental Medicine and Biology

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It has been shown that in rat heart NCX1 exists in a macromolecular -complex including PKA, PKA-anchoring protein, PKC, and phosphatases PP1 and PP2A. In addition, several lines of evidence suggest that the interactions of the exchanger with other molecules are closely associated with its function in regulation of [Ca(2+)](i). NCX contains a large intracellular loop (NCXIL) that is responsible for regulating NCX activity. We used the yeast two-hybrid method to screen a human heart cDNA library and found that the C-terminal region of sarcomeric mitochondrial creatine kinase (sMiCK) interacted with NCX1IL. Among the four creatine kinase (CK) isozymes, both sMiCK and the muscle-type cytosolic creatine kinase (CKM) co-immunoprecipitated with NCX1. Both sMiCK and CKM were able to produce a recovery in the decreased NCX1 activity that was lost under energy-compromised conditions. This regulation is mediated through a putative PKC phosphorylation site of sMiCK and CKM. The catalytic activity of sMiCK and CKM is not required for their regulation of NCX1 activity. Our results suggest a novel mechanism for the regulation of NCX1 activity and a novel role for CK.

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Regulation of muscle creatine kinase by phosphorylation in normal and diabetic hearts

Cited by 25 publications

References 33 publications

Regulation of mitochondrial aconitase by phosphorylation in diabetic rat heart

Regulation of mitochondrial aconitase by phosphorylation in diabetic rat heart

Regulation of Sodium-Calcium Exchanger Activity by Creatine Kinase under Energy-compromised Conditions

Regulation of Sodium-Calcium Exchanger Activity by Creatine Kinase

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