๐Ÿ“’ Garcia 2016

The ATPase Inhibitory Factor 1 (IF1): A master regulator of energy metabolism and of cell survival



  • Mitochondria interact with elements of the cytoskeleton, for their subcellular localization and motility and with the endoplasmic reticulum (ER) via ERMES protein complexes

  • Cristae of the IMM are bound to the OMM by inner membrane peripheral areas known as cristae junctions, depending on mitochondrial contact site and cristae organizing system (MICOS). Cristae locate all complexes of the ETC and thus increase the total surface of the IMM

  • Formation of ATP synthase dimers in a V-shaped structure with an angle of 86ยฐ between monomers generates the curvature on the IMM that promotes cristae formation, melt down with aging.

  • respiratory complexes are assembled in higher order structures named supercomplexes or respirasomes.

    • Increasing efficiency and decreasing ROS formation

The mitochondrial ATP synthase: structure and assembly

  • F1-ATPase domain is extrinsic to the IMM
  • Fo-ATPase domain is embedded in the membrane, a hydrophobic cylindrical structure consisting of a ring of c subunits that along with subunits ฮณ , ฮด and ฮต form the rotor of the engine.
  • A key to catalysis are the conformational changes experienced by the ฮฒ subunits due to the rotation of the ฮณ subunit
  • ATP synthesis needs the proton electrochemical gradient as driving force to power the rotation of the cylinder in the Fodomain while, when functioning in reverse, ATP hydrolysis pumps H+ out to the intermembrane space
  • C5 is also involved in the regulation of cell death: Bax and other apoptosis inducing agents => mPTP

Regulation of the ATP synthase

  • Neclear DNA and mtDNA
  • Transcriptional and post-transcriptional levels
  • activation/ inhibition of the respiratory chain
  • substrate availability
  • covalent modifications of the enzyme
  • the interaction with different regulatory proteins (e.g. IF1)

ATPase Inhibitory Factor 1: gene and protein

  • ATPIF1 gene, three different isoforms by alternative splicing
  • First discovered in bovine heart mitochondria
  • The active form of bovine IF1 is a homodimer

Oligomerization of IF1 and the ATP synthase

  • The optimal inhibitory effect of IF on the hydrolase activity of the enzyme is at an acidic pH as dimers.
  • Rise in pH => forming tetramers and not inhibiting ATP synthase
  • calmodulin (CaM) binding (+): IF1-CaM complex could be regulating the import of IF1 into mitochondria
  • Helping dimerization of ATP synthase as bridges

IF1 binding inhibits both the ATP synthase and hydrolase activities of the ATP synthase

  • Hydrolase-inghibiting function of IF1 is to prevent reverse functioning of the enzyme when mitochondria become de-energized to avoid the wasting of cellular ATP

  • Recovery of ฮ”ฮจ, ATP, and Mg is required for IF1 to leave ATP synthase

  • IF1 in solution, not bound to the ATP synthase, is an intrinsically disordered protein. Phosphorylation events can regulate these transitions. Disordered conformation of the soluble inhibitor is essential for its initial interaction with the F1-ATPase domain of the ATP synthase

Regulation of IF1 expression

  • Expression levels of hypoxia-inducible factor 1 ฮฑ (HIF-1ฮฑ) correlated with the expression levels of IF1 mRNA, but not the whole picture.
  • Most of the evidences regarding the control of IF1 expression point to post-transcriptional regulatory mechanisms, like high accumulation of IF1 observed in carcinomas of the colon, lung, breast and ovarian cancer patients.
  • The immediate early response 3 (IER3) interacts directly with the C-terminal domain of IF1.
  • IF1 is a mitochondrial protein with a very short half-life of ~2โ€“4h, cleaved by mitochondrial serine-proteases

Regulation of IF1 activity

Covalent modifications on IF1

  • NAD+-dependent deacetylase SIRT3 => deacetylation of lysines
  • succinylation?
  • glycosylation
  • phosphorylation

Regulation of IF1 activity by PKA

  • PKA can attach to OMM or inside mitochondria
  • PKA regulates mitochondrial proteins by phosphorylation (e.g. Na+/Ca2+ exchanger)
  • PKA regulates apoptosis, mitochondrial dynamics , mitophagy , metabolism and also oxidative phosphorylation
  • PKA enhances OXPHOS rates, inactivates IF1, increases ATP synthetic capacity of mitochondria
  • Physiologically generated cAMP does not pass through mitochondrial membranes=> depends on an intramitochondrial source of cAMP, soluble adenylyl cyclase (sAC)

Physiologically relevant contexts for the regulation of IF1

Prevention of ATP depletion during hypoxia

  • Reversal of ANT and ATP synthase in ฮ”ฮจ drops. ATP producers => ATP consumers.
  • IF1 is expected to be dephosphorylated and, by binding to the H+-ATP synthase would inhibit the ATP hydrolytic function of the enzyme, delaying ATP depletion, damage and cell death.

Maintenance of the membrane potential/mitophagy

  • Cells overexpressing IF1 in hypoxia had a worse maintenance of the membrane potential than cells with silenced IF1
  • Mitochondrial membrane potential is essential for identifying healthy mitochondria since depolarization triggers the induction of mitophagy, involving PINK1 & Parkin
  • IF1 depleted cells better maintain the membrane potential thus hampering the stabilization of PINK1 and the recruitment of Parkin.

Metabolic reprogramming in cancer and in cellular differentiation

  • Cancer cells down-regulate OXPHOS and tactivate aerobic glycolysis, optimal for cell proliferation.
  • silencing of OXPHOS in cancer is also exerted by the overexpression of IF1
  • inhibition of the ATP synthase by overexpression of IF1 promotes mitochondrial hyperpolarization, increasing ROS generation. Activation of different survival pathways by inducing a state of pre-conditioning = mitohormesis
  • IF1 by potentially regulating the oligomeric state of the ATP synthase might contribute to stabilize cristae structure hence upgrading, at the structural level, the threshold for cell death.
  • high expression level of IF1 in the tumor is a bad predictor of survival and recurrence of the disease in liver, bladder, gastric and glioma cancer patient, but better in breast and colon carcinomas


  1. Garcรญa-Bermรบdez J, Cuezva JM. The ATPase Inhibitory Factor 1 (IF1): A master regulator of energy metabolism and of cell survival. Biochim. Biophys. Acta 2016;1857(8):1167-1182. doi:10.1016/j.bbabio.2016.02.004. ↩︎