Accessory subunit NDUFB4 participates in mitochondrial complex I supercomplex formation
Gaganvir Parmar, Claire Fong-McMaster, Chantal A. Pileggi, David A. Patten, Alexanne Cuillerier, Stephanie Myers, Ying Wang, Siegfried Hekimi, Miroslava Čuperlović‐Culf, Mary‐Ellen Harper
Abstract
Mitochondrial electron transport chain (ETC) complexes organize into supramolecular structures called respiratory supercomplexes (SCs). The role of respiratory SC remains largely unconfirmed despite evidence supporting their necessity for mitochondrial respiratory function. The mechanisms underlying the formation of the I 1 III 2 IV 1 "respirasome" SC are also not fully understood, further limiting insights into these processes in physiology and diseases, including neurodegeneration and metabolic syndromes. NDUFB4 is a complex I accessory subunit that contains residues that interact with the subunit UQCRC1 from complex III, suggesting that NDUFB4 is integral for I 1 III 2 IV 1 respirasome integrity. Here, we introduced specific point mutations to Asn24 (N24) and Arg30 (R30) residues on NDUFB4 to decipher the role of I 1 III 2 -containing respiratory SCs in cellular metabolism while minimizing the functional consequences to complex I assembly. Our results demonstrate that NDUFB4 point mutations N24A and R30A impair I 1 III 2 IV 1 respirasome assembly and reduce mitochondrial respiratory flux. Steady-state metabolomics also revealed a global decrease in TCA cycle metabolites, affecting NADH-generating substrates. Taken together, our findings highlight an integral role of NDUFB4 in respirasome assembly and demonstrate the functional significance of SCs in regulating mammalian cell bioenergetics.