Abstract
Aggressive cancers exhibit an efficient conversion of high amounts of glucose to lactate accompanied by acid secretion, a phenomenon popularly known as the Warburg effect. The acidic microenvironment and the alkaline cytosol create a proton-gradient (acid gradient) across the plasma membrane that represents proton-motive energy. Increasing experimental data from physiological relevant models suggest that acid gradient stimulates tumor proliferation, and can also support its energy needs. However, direct biochemical evidence linking extracellular acid gradient to generation of intracellular ATP are missing. In this work, we demonstrate that cancer cells can synthesize significant amounts of phosphate-bonds from phosphate in response to acid gradient across plasma membrane. The noted phenomenon exists in absence of glycolysis and mitochondrial ATP synthesis, and is unique to cancer. Biochemical assays using viable cancer cells, and purified plasma membrane vesicles utilizing radioactive phosphate, confirmed phosphate-bond synthesis from free phosphate (Pi), and also localization of this activity to the plasma membrane. In addition to ATP, predominant formation of pyrophosphate (PPi) from Pi was also observed when plasma membrane vesicles from cancer cells were subjected to trans-membrane acid gradient. Cancer cytosols were found capable of converting PPi to ATP, and also stimulate ATP synthesis from Pi from the vesicles. Acid gradient created through glucose metabolism by cancer cells, as observed in tumors, also proved critical for phosphate-bond synthesis. In brief, these observations reveal a role of acidic tumor milieu as a potential energy source and may offer a novel therapeutic target.
Highlights
Warburg effect is a metabolic hallmark of most aggressive cancer cells whereby most of the glucose is converted to lactate in presence of oxygen [1, 2]
We designed an experiment to simultaneously measure the levels of ATP, lactate, glucose and the pH of the external medium in real time during aerobic glycolysis of highly glycolytic breast cancer cells, MDA-MB-231[24]
The rates of glucose consumption (3.46 nmol/min/million-cells, s.d. = ±0.17, 3 data sets) and lactate production (5.95 nmol/ min/million-cells, s.d. = ±0.50, 3 data sets) exhibited no significant change (Fig 1D). This experiment indicated that the extracellular acid gradient plays a critical role even during aerobic glycolysis in maintaining high levels of ATP in cancer cells
Summary
Warburg effect is a metabolic hallmark of most aggressive cancer cells whereby most of the glucose is converted to lactate in presence of oxygen [1, 2]. The aerobic glycolysis is accompanied by acidification of the tumor microenvironment [3, 4] that confers selective growth advantage to cancer cells by metabolic reprogramming [5,6,7], increased invasiveness [8,9,10] and regulation of cell cycle [11]. Even though aerobic glycolysis and extracellular acidification confers selective growth advantage to cancer as discussed above, as to how the cancer cells meet its energy demand under this condition remains a paradox. The cellular machinery subsequently utilizes this energy to drive the synthesis of high-energy phosphate bonds in the form of ATP and other high-energy molecules for utilization in cellular processes. [13, 16]
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