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Integration of flux measurements to resolve changes in anabolic and catabolic metabolism in cardiac myocytes
Although ancillary pathways of glucose metabolism are critical for synthesizing cellular building blocks and modulating stress responses, how they are regulated remains unclear. In the present study, we used radiometric glycolysis assays, [(13)C(6)]-glucose isotope tracing, and extracellular flux an...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Portland Press Ltd.
2017
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5545928/ https://www.ncbi.nlm.nih.gov/pubmed/28706006 http://dx.doi.org/10.1042/BCJ20170474 |
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author | Gibb, Andrew A. Lorkiewicz, Pawel K. Zheng, Yu-Ting Zhang, Xiang Bhatnagar, Aruni Jones, Steven P. Hill, Bradford G. |
author_facet | Gibb, Andrew A. Lorkiewicz, Pawel K. Zheng, Yu-Ting Zhang, Xiang Bhatnagar, Aruni Jones, Steven P. Hill, Bradford G. |
author_sort | Gibb, Andrew A. |
collection | PubMed |
description | Although ancillary pathways of glucose metabolism are critical for synthesizing cellular building blocks and modulating stress responses, how they are regulated remains unclear. In the present study, we used radiometric glycolysis assays, [(13)C(6)]-glucose isotope tracing, and extracellular flux analysis to understand how phosphofructokinase (PFK)-mediated changes in glycolysis regulate glucose carbon partitioning into catabolic and anabolic pathways. Expression of kinase-deficient or phosphatase-deficient 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase in rat neonatal cardiomyocytes co-ordinately regulated glycolytic rate and lactate production. Nevertheless, in all groups, >40% of glucose consumed by the cells was unaccounted for via catabolism to pyruvate, which suggests entry of glucose carbons into ancillary pathways branching from metabolites formed in the preparatory phase of glycolysis. Analysis of (13)C fractional enrichment patterns suggests that PFK activity regulates glucose carbon incorporation directly into the ribose and the glycerol moieties of purines and phospholipids, respectively. Pyrimidines, UDP-N-acetylhexosamine, and the fatty acyl chains of phosphatidylinositol and triglycerides showed lower (13)C incorporation under conditions of high PFK activity; the isotopologue (13)C enrichment pattern of each metabolite indicated limitations in mitochondria-engendered aspartate, acetyl CoA and fatty acids. Consistent with this notion, high glycolytic rate diminished mitochondrial activity and the coupling of glycolysis to glucose oxidation. These findings suggest that a major portion of intracellular glucose in cardiac myocytes is apportioned for ancillary biosynthetic reactions and that PFK co-ordinates the activities of the pentose phosphate, hexosamine biosynthetic, and glycerolipid synthesis pathways by directly modulating glycolytic intermediate entry into auxiliary glucose metabolism pathways and by indirectly regulating mitochondrial cataplerosis. |
format | Online Article Text |
id | pubmed-5545928 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2017 |
publisher | Portland Press Ltd. |
record_format | MEDLINE/PubMed |
spelling | pubmed-55459282017-08-16 Integration of flux measurements to resolve changes in anabolic and catabolic metabolism in cardiac myocytes Gibb, Andrew A. Lorkiewicz, Pawel K. Zheng, Yu-Ting Zhang, Xiang Bhatnagar, Aruni Jones, Steven P. Hill, Bradford G. Biochem J Research Articles Although ancillary pathways of glucose metabolism are critical for synthesizing cellular building blocks and modulating stress responses, how they are regulated remains unclear. In the present study, we used radiometric glycolysis assays, [(13)C(6)]-glucose isotope tracing, and extracellular flux analysis to understand how phosphofructokinase (PFK)-mediated changes in glycolysis regulate glucose carbon partitioning into catabolic and anabolic pathways. Expression of kinase-deficient or phosphatase-deficient 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase in rat neonatal cardiomyocytes co-ordinately regulated glycolytic rate and lactate production. Nevertheless, in all groups, >40% of glucose consumed by the cells was unaccounted for via catabolism to pyruvate, which suggests entry of glucose carbons into ancillary pathways branching from metabolites formed in the preparatory phase of glycolysis. Analysis of (13)C fractional enrichment patterns suggests that PFK activity regulates glucose carbon incorporation directly into the ribose and the glycerol moieties of purines and phospholipids, respectively. Pyrimidines, UDP-N-acetylhexosamine, and the fatty acyl chains of phosphatidylinositol and triglycerides showed lower (13)C incorporation under conditions of high PFK activity; the isotopologue (13)C enrichment pattern of each metabolite indicated limitations in mitochondria-engendered aspartate, acetyl CoA and fatty acids. Consistent with this notion, high glycolytic rate diminished mitochondrial activity and the coupling of glycolysis to glucose oxidation. These findings suggest that a major portion of intracellular glucose in cardiac myocytes is apportioned for ancillary biosynthetic reactions and that PFK co-ordinates the activities of the pentose phosphate, hexosamine biosynthetic, and glycerolipid synthesis pathways by directly modulating glycolytic intermediate entry into auxiliary glucose metabolism pathways and by indirectly regulating mitochondrial cataplerosis. Portland Press Ltd. 2017-08-15 2017-08-07 /pmc/articles/PMC5545928/ /pubmed/28706006 http://dx.doi.org/10.1042/BCJ20170474 Text en © 2017 The Author(s) https://creativecommons.org/licenses/by-nc-nd/4.0/This is an open access article published by Portland Press Limited on behalf of the Biochemical Society and distributed under the Creative Commons Attribution License 4.0 (CC BY-NC-ND) (https://creativecommons.org/licenses/by-nc-nd/4.0/) . |
spellingShingle | Research Articles Gibb, Andrew A. Lorkiewicz, Pawel K. Zheng, Yu-Ting Zhang, Xiang Bhatnagar, Aruni Jones, Steven P. Hill, Bradford G. Integration of flux measurements to resolve changes in anabolic and catabolic metabolism in cardiac myocytes |
title | Integration of flux measurements to resolve changes in anabolic and catabolic metabolism in cardiac myocytes |
title_full | Integration of flux measurements to resolve changes in anabolic and catabolic metabolism in cardiac myocytes |
title_fullStr | Integration of flux measurements to resolve changes in anabolic and catabolic metabolism in cardiac myocytes |
title_full_unstemmed | Integration of flux measurements to resolve changes in anabolic and catabolic metabolism in cardiac myocytes |
title_short | Integration of flux measurements to resolve changes in anabolic and catabolic metabolism in cardiac myocytes |
title_sort | integration of flux measurements to resolve changes in anabolic and catabolic metabolism in cardiac myocytes |
topic | Research Articles |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5545928/ https://www.ncbi.nlm.nih.gov/pubmed/28706006 http://dx.doi.org/10.1042/BCJ20170474 |
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