Tumour suppressor p53 controls biosynthesis through inactivating the key enzyme glucose-6-phosphate dehydrogenase

The tumour suppressor gene p53 is the most commonly mutated gene in human cancer. Its role in normal cells is to regulate the cell cycle and prevent DNA mutation, hence its common tag of “guardian of the genome”. However, the complete range of its anti-proliferative functions is not yet wholly understood.

Cancer cells produce energy through glycolysis even when oxygen levels are high; this so-called “Warburg effect” is thought to drive them to divert energy into biosynthesis and thus to both grow and divide rapidly. It has recently been suggested that p53 might have a role in regulating glycolysis, and thence ATP production and biosynthesis, through the pentose phosphate pathway of oxidative phosphorylation (PPP). Mian Wu of the University of Science and Technology of China, Hefei, China, Xiaolu Yang of the University of Pennsylvania School of Medicine, Philadelphia, PA, USA and their co-workers have now explored this putative link between p53 and the pentose phosphate pathway in human cancer cells.

HCT116 colon cancer cells with wild type p53 (p53^+/+) and identical cells with p53 deleted (p53^-/-) were cultured in a medium containing ¹³C glucose, and the glucose metabolites measured using NMR spectroscopy. Glucose metabolism through the PPP pathway was increased by about 50% in the p53^-/- cells. As expected, this led to an increase in NADPH levels in these cells. This effect was also seen in wild type cells in which p53 expression had been knocked down using a small hairpin RNA (shRNA); however, the effect could be almost completely reversed by down-regulation of the first enzyme in this pathway, glucose-6-phosphate dehydrogenase (G6PD). To verify the result further, these researchers compared NADPH levels in different tissues in wild type and p53^-/- mice, and found that p53 depletion increased glucose metabolism and NADPH levels in all tissues other than the spleen.

NADPH is known to be required for lipid metabolism. The researchers tested the effect of p53 depletion on this by treating p53^+/+ and p53^-/- cancer cells with a combination of drugs that stimulate lipogenesis. They found an increase in lipid production in the p53^-/- cells that, similarly, could be reduced by blocking G6PD. Levels of G6PD expression were unchanged in the p53^-/- cells in all lines tested, however, indicating that p53 suppresses G6PD activity but not its expression.

The researchers then showed, using a variety of biophysical techniques, that wild type p53 interacts directly with G6PD; that this interaction takes place in the cytoplasm; and that the interaction occurs in vitro and in yeast and E. coli cells, where p53 binding suppresses G6PD activity. A panel of mutants in which different p53 domains were deleted was used to determine the parts of this complex protein that are necessary for both G6PD binding and inhibition. This was shown to involve four separate p53 domains, two within the protein’s C-terminal region. Many variants of p53 containing tumour-associated missense mutations were found to be unable to inhibit G6PD, although some of these still bound the enzyme.

In vivo, G6PD exists in equilibrium between a monomer and a dimer, the latter being its active form. The researchers found that dimer levels decreased when active p53 was present, which strongly suggests that p53 inhibits G6PD activity by preventing dimerisation. This enzyme is the rate-limiting step of the pentose phosphate pathway, which is essential for biosynthesis. Wu, Yang and their colleagues concluded that suppressing this pathway by preventing G6PD dimerisation is a key role for p53 in normal cells, and that p53-inactivating mutations may promote G6PD activity and thus biosynthesis in tumour cells.

Reference

Jiang, P., Du1, W., Wang, X., Mancuso, A., Gao, X,. Wul. M. and Yang, X. (2011). p53 regulates biosynthesis through direct inactivation of glucose-6-phosphate dehydrogenase. Nature Cell Biology, published online ahead of print 20 February 2011. DOI: 10.1038/ncb2172