Interaction between AIF and CHCHD4 Regulates Respiratory Chain Biogenesis

Abstract

Apoptosis-inducing factor (AIF) is a mitochondrial flavoprotein that, beyond its apoptotic function, is required for the normal expression of major respiratory chain complexes. Here we identified an AIF-interacting protein, CHCHD4, which is the central component of a redox-sensitive mitochondrial intermembrane space import machinery. Depletion or hypomorphic mutation of AIF caused a downregulation of CHCHD4 protein by diminishing its mitochondrial import. CHCHD4 depletion sufficed to induce a respiratory defect that mimicked that observed in AIF-deficient cells. CHCHD4 levels could be restored in AIF-deficient cells by enforcing its AIF-independent mitochondrial localization. This modified CHCHD4 protein reestablished respiratory function in AIF-deficient cells and enabled AIF-deficient embryoid bodies to undergo cavitation, a process of programmed cell death required for embryonic morphogenesis. These findings explain how AIF contributes to the biogenesis of respiratory chain complexes, and they establish an unexpected link between the vital function of AIF and the propensity of cells to undergo apoptosis.

Mitochondrial Proteins Containing Coiled-Coil-Helix-Coiled-Coil-Helix (CHCH) Domains in Health and Disease

“…Even though the principal catalytic and structural features of Mia40, in particular within the 94 segment that contains the redox active cysteine-proline-cysteine (CPC) and the (CX 9 C) 2 95 motifs, have been conserved throughout eukaryotic evolution [27, [36][37][38][39][40], there are important 96 differences between yeast and mammals. In contrast to yeast Mia40 (~40 kDa), human 97 CHCHD4 does not possess an N-terminal presequence and hence lacks a membrane anchor 98…”

“…[27, [36][37][38]40]. Thus, CHCHD4 is a relatively small (16 kDa) soluble protein that is no 99 longer imported via the TIM23 translocase but rather relies on a self-catalytic import process 100 [36,37].…”

“…This switch in the import pathway might have occurred during evolution to couple 101 the import of CHCHD4 more tightly to the IMS redox-signaling pathway, perhaps to achieve 102 rapid adaptive responses in mitochondria responding to ever changing metabolic needs. 103 CHCHD4 has lost its hydrophobic inner membrane anchor, yet has acquired the capacity to 104 interact with a new partner, the flavoprotein AIF [40], which is attached to the inner 105 membrane [41,42]. The N-terminal 27 residues of CHCHD4 are necessary and sufficient to 106 establish a direct interaction with AIF [40].…”

“…103 CHCHD4 has lost its hydrophobic inner membrane anchor, yet has acquired the capacity to 104 interact with a new partner, the flavoprotein AIF [40], which is attached to the inner 105 membrane [41,42]. The N-terminal 27 residues of CHCHD4 are necessary and sufficient to 106 establish a direct interaction with AIF [40]. Nuclear magnetic resonance (NMR) spectroscopy 107…”

“…However, far-UV circular dichroism 109 (CD) spectroscopy analysis suggests that upon its interaction with AIF, the AIF-binding 110 segment of CHCHD4 undergoes important conformational rearrangements that may well 111 affect the interactions of the CHCHD4 CPC domain with its incoming substrates or its 112 reoxidizing partner ERV1/ALR [40]. The conformational status of AIF, which is determined 113 by the binding of its cofactor NADH [43], appears crucial for its interaction with CHCHD4.…”