The possible dual role of IF1 in tumor metabolism: its contribution to the Warburg effect and promotion of anoxic tumor cell proliferation after re-oxygenation

Tumour cells exhibit high expression of IF1, an endogenous protein that inhibits the hydrolytic activity of ATP synthase under conditions of reduced mitochondrial membrane potential (∆μH+), as occurs in ischaemia. The pro-oncogenic effects of the inhibitor include the reprogramming of cellular energy metabolism towards the Warburg effect, which is promoted by IF1 in the dephosphorylated state. Both metabolic reprogramming and its PKA-dependent phosphorylation are highly controversial. To clarify these critical points, stable clones with silenced IF1 were generated and compared with parental IF1-expressing tumour cell lines. Bioenergetic parameters such as respiration rate, ATP synthesis rate (OXPHOS) and mitochondrial membrane potential were similar in IF1-silenced and control cells. This clearly shows that IF1 does not inhibit ATP synthase in tumour cells when the enzyme functions physiologically. Furthermore, cells showed a similar OXPHOS rate when exposed to positive and negative PKA modulators and stimulated with NAD+ or FAD-dependent substrates, regardless of the presence or absence of IF1. Consequently, the results of this work exclude the dependence of the effect of IF1 on its phosphorylated/dephosphorylated state. Our study clearly shows that IF1 does not inhibit mitochondrial ATP synthase or OXPHOS in normoxic tumour cells, thus invalidating its putative contribution to the Warburg effect. The role of IF1 as a pro-oncogenic factor is to protect tumour cells from apoptosis and severe hypoxia. We have investigated its influence on cancer cell survival and proliferation under conditions mimicking ischaemia-reperfusion. Experiments with IF1-silenced and parental cells exposed to the FCCP uncoupler showed that mitochondrial mass was preserved in all uncoupled cells. IF1-expressing cells exhibited increased mitophagy, which was balanced by mitochondrial biogenesis, resulting in a higher energy charge and a proliferative advantage upon re-oxygenation. IF1 proves to be a promising therapeutic target against OXPHOS-dependent tumours.