C subunit of the ATP synthase is an amyloidogenic calcium dependent channel-forming peptide with possible implications in mitochondrial permeability transition

Abstract

The c subunit is an inner mitochondrial membrane (IMM) protein encoded by three nuclear genes. Best known as an integral part of the F0 complex of the ATP synthase, the c subunit is also present in other cytoplasmic compartments in ceroid lipofuscinoses. Under physiological conditions, this 75 residue-long peptide folds into an α-helical hairpin and forms oligomers spanning the lipid bilayer. In addition to its physiological role, the c subunit has been proposed as a key participant in stress-induced IMM permeabilization by the mechanism of calcium-induced permeability transition. However, the molecular mechanism of the c subunit participation in IMM permeabilization is not completely understood. Here we used fluorescence spectroscopy, atomic force microscopy and black lipid membrane methods to gain insights into the structural and functional properties of unmodified c subunit protein that might make it relevant to mitochondrial toxicity. We discovered that c subunit is an amyloidogenic peptide that can spontaneously fold into β-sheets and self-assemble into fibrils and oligomers in a Ca2+-dependent manner. C subunit oligomers exhibited ion channel activity in lipid membranes. We propose that the toxic effects of c subunit might be linked to its amyloidogenic properties and are driven by mechanisms similar to those of neurodegenerative polypeptides such as Aβ and α-synuclein.

Formation of High-Conductive C Subunit Channels upon Interaction with Cyclophilin D

“…Figure 1A shows a representative trace of the c subunit oligomers at 50 mV. This channel activity is in good agreement with previously reported activity of the unmodified peptide [8]. In order to study the effect of CypD on the electrophysiological activity of the c subunit, we dissolved the lyophilized peptide in the same buffer as above, but this time, it was supplemented with 0.41 mg/mL of CypD.…”

“…This mechanism is supported by studies that showed that the c subunit alone is sufficient to induce PTP [6,12,14]. Our recent report showed that the unmodified c subunit is an amyloidogenic peptide that spontaneously folds into cross-β oligomers and offers a potential mechanism on how such pores can form [8]. One of the defining features of the channel candidate for involvement in PTP is its dependence on CypD, an endogenous mitochondrial chaperone and well-established activator of PTP [16].…”

“…Other studies suggest a separate mechanism where amyloidogenic peptides are involved in the formation of PTP in aging-related disorders. Interestingly, we discovered that the c subunit of the ATP synthase is in fact an amyloidogenic peptide [8,9]. Despite PTP ion conducting pores formed by these protein complexes exhibit different conductances, their involvement in the channel opening depends on the interaction with cyclophilin D (CypD), a mitochondrial chaperone and the specific target of cyclosporin A (CSA) [10].…”

Protein folding and unfolding: proline cis - trans isomerization at the c subunits of F 1 F O -ATPase might open a high conductance ion channel

“…The CsA can inhibit the mPTP opening at an early stage but not at later ones. The c subunits could also form an ion channel by assembling into oligomers in a β-sheet conformation with a similar mechanism to some other amyloidogenic peptides that form a β-sheet oligomeric pore [19]. Recently, it has been suggested that CyPD-c subunit interactions help the formation of higher-order oligomers, but is not required for pore activity by highlighting the folding activity in the mPTP formation [20].…”

From the Structural and (Dys)function of ATP Synthase to Deficiency in Age-Related Diseases

“…Several proteins such as VIDAC, ANT and CypD have been considered a potential forming channel but robust results are for ATP synthase dimers or its c-ring, the major component of the membrane FO complex of the enzyme, as the core channel of mPTP [50,101,102]. Indeed, high Ca 2+ concentration inhibits the organization of c subunit in fibrils and induces the formation of cross-β oligomers, able to form small channels in model lipid bilayers [50,101,101,103,104].…”