The pyruvate dehydrogenase complex (PDC) is a multienzyme complex that plays a key role in energy metabolism by converting pyruvate to acetyl-CoA. An increase of nuclear PDC has been shown to be correlated with an increase of histone acetylation that requires acetyl-CoA.PDC has been reported to form a ~ 10 MDa macromolecular machine that is proficient in performing sequential catalytic reactions via its three components. In this study, we show that the PDC displays size versatility in an ionic strength-dependent manner using size exclusion chromatography of yeast cell extracts. Biochemical analysis in combination with mass spectrometry indicates that yeast PDC (yPDC) is a salt-labile complex that dissociates into submegadalton individual components even under physiological ionic strength. Interestingly, we find that each oligomeric component of yPDC displays a larger size than previously believed. In addition, we show that the mammalian PDC also displays this uncommon characteristic of saltlability, although it has a somewhat different profile compared to yeast. We show that the activity of yPDC is reduced in higher ionic strength. Our results indicate that the structure of PDC may not always maintain its ~ 10 MDa organization, but is rather variable. We propose that the flexible nature of PDC may allow modulation of its activity.Keywords: PDC, mitochondrial metabolism, TCA cycle, pyruvate, acetyl-CoA, highperformance liquid chromatography (HPLC), size exclusion chromatography (SEC) synthesis, needs PDC to convert pyruvate into acetyl-CoA [1,2]. Being a key enzyme in cellular metabolism, PDC is also considered a target for anticancer and antibacterial drugs [3,4].Furthermore, the level of nuclear PDC is correlated with that of histone acetylation as well as its recruitment to some promoters upon induction possibly for local acetyl-CoA production [5][6][7][8].The central role of PDC in energy homeostasis necessitates a tight regulation of its activity. Short-term regulation (minutes to hours) occurs through the inhibitory phosphorylation of E1α by pyruvate dehydrogenase kinases (PDKs) and phosphatases, while long-term regulation of PDC activity can be exerted at transcriptional levels within days to weeks [9,10]. PDKs have been studied as potential drug targets as well because of their upregulation or activation in diseases such as diabetes and cancer [11,12]. The catalysis of PDC is performed by three separate enzymes E1p (pyruvate decarboxylase), E2p (dihydrolipoamide acyltransferase), and E3 (dihydrolipoamide dehydrogenase), which are linked together efficiently into a large multienzyme complex, where the oligomeric E2p forms a structural core, to which multiple copies of the E1p, E3, and E3BP (E3 binding protein) are bound [13]. The E1p performs the rate-limiting oxidative decarboxylation of pyruvate via thiamine pyrophosphate (TPP) and transfers the acetyl-group to the lipoyl group of the E2p. Then the E2p transfers the acyl group to CoA while reducing its lipoyl domain, which is then oxidized by the E3 which...
Changes in available nutrients are inevitable events for most living organisms. Upon nutritional stress, several signaling pathways cooperate to change the transcription program through chromatin regulation to rewire cellular metabolism. In budding yeast, histone H3 threonine 11 phosphorylation (H3pT11) acts as a marker of low glucose stress and regulates the transcription of nutritional stress-responsive genes. Understanding how this histone modification ‘senses’ external glucose changes remains elusive. Here, we show that Tda1, the yeast ortholog of human Nuak1, is a direct kinase for H3pT11 upon low glucose stress. Yeast AMP-activated protein kinase (AMPK) directly phosphorylates Tda1 to govern Tda1 activity, while CK2 regulates Tda1 nuclear localization. Collectively, AMPK and CK2 signaling converge on histone kinase Tda1 to link external low glucose stress to chromatin regulation.
The pyruvate dehydrogenase complex (PDC) is a multienzyme complex that plays a key role in energy metabolism by converting pyruvate to acetyl-CoA. An increase of nuclear PDC has been shown to be correlated with an increase of histone acetylation that requires acetyl-CoA. PDC has been reported to form a ~ 10 MDa macromolecular machine that is proficient in performing sequential catalytic reactions via its three components. In this study, we show that the PDC displays size versatility in an ionic strength-dependent manner using size exclusion chromatography of yeast cell extracts. Biochemical analysis in combination with mass spectrometry indicates that yeast PDC (yPDC) is a salt-labile complex that dissociates into sub-megadalton individual components even under physiological ionic strength. Interestingly, we find that each oligomeric component of yPDC displays a larger size than previously believed. In addition, we show that the mammalian PDC also displays this uncommon characteristic of salt-lability, although it has a somewhat different profile compared to yeast. We show that the activity of yPDC is reduced in higher ionic strength. Our results indicate that the structure of PDC may not always maintain its ~ 10 MDa organization, but is rather variable. We propose that the flexible nature of PDC may allow modulation of its activity.
No abstract
Changes in available nutrients are inevitable events for most living organisms. Upon nutritional stress, several signaling pathways cooperate to change the transcription program through chromatin regulation to rewire cellular metabolism. In budding yeast, histone H3 threonine 11 phosphorylation (H3pT11) acts as a marker of low glucose stress and regulates the transcription of nutritional stress responsive genes. Understanding how this histone modification senses external glucose changes remains elusive. Here, we show that Tda1, the yeast orthologue of human Nuak1, is a direct kinase for H3pT11 upon low glucose stress. Yeast AMPK directly phosphorylates Tda1 to govern Tda1 activity, while CK2 regulates Tda1 nuclear localization. Collectively, AMPK and CK2 signaling converge on histone kinase Tda1 to link external low glucose stress to chromatin regulation.
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