In the last two decades, IF 1 , the endogenous inhibitor of the mitochondrial F 1 F o -ATPase (ATP synthase) has assumed greater and ever greater interest since it has been found to be overexpressed in many cancers. At present, several findings indicate that IF 1 is capable of playing a central role in cancer cells by promoting metabolic reprogramming, proliferation and resistance to cell death. However, the mechanism(s) at the basis of this pro-oncogenic action of IF 1 remains elusive. Here, we recall the main features of the mechanism of the action of IF 1 when the ATP synthase works in reverse, and discuss the experimental evidence that support its relevance in cancer cells. In particular, a clear pro-oncogenic action of IF 1 is to avoid wasting of ATP when cancer cells are exposed to anoxia or near anoxia conditions, therefore favoring cell survival and tumor growth. However, more recently, various papers have described IF 1 as an inhibitor of the ATP synthase when it is working physiologically (i.e. synthethizing ATP), and therefore reprogramming cell metabolism to aerobic glycolysis. In contrast, other studies excluded IF 1 as an inhibitor of ATP synthase under normoxia, providing the basis for a hot debate. This review focuses on the role of IF 1 as a modulator of the ATP synthase in normoxic cancer cells with the awareness that the knowledge of the molecular action of IF 1 on the ATP synthase is crucial in unravelling the molecular mechanism(s) responsible for the pro-oncogenic role of IF 1 in cancer and in developing related anticancer strategies.

The ATP synthase (F o F 1 ) of Escherichia coli couples the translocation of protons across the cytoplasmic membrane by F o to ATP synthesis or hydrolysis in F 1 . Whereas good knowledge of the nanostructure and the rotary mechanism of the ATP synthase is at hand, the assembly pathway of the 22 polypeptide chains present in a stoichiometry of ab 2 c 10 α 3 β 3 γδϵ has so far not received sufficient attention. In our studies, mutants that synthesize different sets of F o F 1 subunits allowed the characterization of individually formed stable subcomplexes. Furthermore, the development of a time-delayed in vivo assembly system enabled the subsequent synthesis of particular missing subunits to allow the formation of functional ATP synthase complexes. These observations form the basis for a model that describes the assembly pathway of the E. coli ATP synthase from pre-formed subcomplexes, thereby avoiding membrane proton permeability by a concomitant assembly of the open H + -translocating unit within a coupled F o F 1 complex.

The ATP synthases are multiprotein complexes found in the energy-transducing membranes of bacteria, chloroplasts and mitochondria. They employ a transmembrane protonmotive force, Δ p , as a source of energy to drive a mechanical rotary mechanism that leads to the chemical synthesis of ATP from ADP and P i . Their overall architecture, organization and mechanistic principles are mostly well established, but other features are less well understood. For example, ATP synthases from bacteria, mitochondria and chloroplasts differ in the mechanisms of regulation of their activity, and the molecular bases of these different mechanisms and their physiological roles are only just beginning to emerge. Another crucial feature lacking a molecular description is how rotation driven by Δ p is generated, and how rotation transmits energy into the catalytic sites of the enzyme to produce the stepping action during rotation. One surprising and incompletely explained deduction based on the symmetries of c-rings in the rotor of the enzyme is that the amount of energy required by the ATP synthase to make an ATP molecule does not have a universal value. ATP synthases from multicellular organisms require the least energy, whereas the energy required to make an ATP molecule in unicellular organisms and chloroplasts is higher, and a range of values has been calculated. Finally, evidence is growing for other roles of ATP synthases in the inner membranes of mitochondria. Here the enzymes form supermolecular complexes, possibly with specific lipids, and these complexes probably contribute to, or even determine, the formation of the cristae.

Methanogenic archaea live at the thermodynamic limit of life and use sophisticated mechanisms for ATP synthesis and energy coupling. The group of methanogens without cytochromes use an Na + current across the membrane for ATP synthesis, whereas the cytochrome-containing methanogens have additional coupling sites that also translocate protons. The ATP synthase in this group is promiscuous and uses Na + and H + simultaneously.

F-type H + -ATP synthases synthesize ATP from ADP and phosphate using the energy supplied by a transmembrane electrochemical potential difference of protons. Rotary subunit movements within the enzyme drive catalysis in either an ATP hydrolysis or an ATP synthesis direction respectively. To monitor these subunit movements and associated conformational changes in real time and with subnanometre resolution, a single-molecule FRET (fluorescence resonance energy transfer) approach has been developed using the double-labelled H + -ATP synthase from Escherichia coli . After reconstitution into a liposome, this enzyme was able to catalyse ATP synthesis when the membrane was energized.