Klepinin 2016 J Bioenerg Biomembr

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Klepinin A, Ounpuu L, Guzun R, Chekulayev V, Timohhina N, Tepp K, Shevchuk I, Schlattner U, Kaambre T (2016) Simple oxygraphic analysis for the presence of adenylate kinase 1 and 2 in normal and tumor cells. J Bioenerg Biomembr 48:531-48.

» PMID: 27854030

Klepinin A, Ounpuu L, Guzun R, Chekulayev V, Timohhina N, Tepp K, Shevchuk I, Schlattner U, Kaambre T (2016) J Bioenerg Biomembr

Abstract: The adenylate kinase (AK) isoforms network plays an important role in the intracellular energy transfer processes, the maintenance of energy homeostasis, and it is a major player in AMP metabolic signaling circuits in some highly-differentiated cells. For this purpose, a rapid and sensitive method was developed that enables to estimate directly and semi-quantitatively the distribution between cytosolic AK1 and mitochondrial AK2 localized in the intermembrane space, both in isolated cells and tissue samples (biopsy material). Experiments were performed on isolated rat mitochondria or permeabilized material, including undifferentiated and differentiated neuroblastoma Neuro-2a cells, HL-1 cells, isolated rat heart cardiomyocytes as well as on human breast cancer postoperative samples. In these samples, the presence of AK1 and AK2 could be detected by high-resolution respirometry due to the functional coupling of these enzymes with ATP synthesis. By eliminating extra-mitochondrial ADP with an excess of pyruvate kinase and its substrate phosphoenolpyruvate, the coupling of the AK reaction with mitochondrial ATP synthesis could be quantified for total AK and mitochondrial AK2 as a specific AK index. In contrast to the creatine kinase pathway, the AK phosphotransfer pathway is up-regulated in murine neuroblastoma and HL-1 sarcoma cells and in these malignant cells expression of AK2 is higher than AK1. Differentiated Neuro-2a neuroblastoma cells exhibited considerably higher OXPHOS capacity than undifferentiated cells, and this was associated with a remarkable decrease in their AK activity. The respirometric method also revealed a considerable difference in mitochondrial affinity for AMP between non-transformed cells and tumor cells.

Keywords: Adenylate kinase, Breast cancer, Cardiomyocytes, Mitochondria, Neuroblastoma, Mouse neuroblastoma Neuro-2A cells, Mouse cardiac sarcoma HL-1 cell

O2k-Network Lab: EE Tallinn Kaambre T, EE Tallinn Kaambre T, EE Tallinn Saks VA, FR Grenoble Schlattner U

Labels: MiParea: Respiration, Comparative MiP;environmental MiP  Pathology: Cancer 

Organism: Human, Mouse, Rat  Tissue;cell: Heart, Nervous system, Endothelial;epithelial;mesothelial cell, Genital  Preparation: Permeabilized cells, Permeabilized tissue, Isolated mitochondria 

Regulation: ATP, AMP  Coupling state: OXPHOS  Pathway: N, NS  HRR: Oxygraph-2k