HiRIEF Proteomics of Arabidopsis thaliana Col-0 and CIAF1 mutants

The assembly of the mitochondrial Complex I requires the expression of nuclear and mitochondrial located genes, co-factor biosynthesis and the assembly of at least 44 subunits. This requires the involvement of assembly factors that interact with subunits of Complex I but are not part of the final holocomplex. A novel plant specific mitochondrial matrix protein encoded by At1g76060, named COMPLEX I ASSEMBLY FACTOR 1 (CIAF1), containing a conserved LYR domain, was shown to be required for Complex I activity. T-DNA insertion mutants of At1g76060 lack the monomeric Complex I and the Supercomplex I+III, displaying the common Complex I growth deficient phenotype. Assembly of Complex I is stalled at 650 and 800 kDa intermediates in mitochondria isolated from these mutants. Evidence points to CIAF1 playing an essential role in the assembly of the peripheral matrix arm Complex I subunits to form the holoenzyme Complex I. We here performed quantitative proteomics by HiRIEF LC-MS with a TMT10plex isobaric tag labeling strategy comparing the control plant Col-0 and several mutant lines, including the mutant with inactivating insertion in At1g76060 (a.k.a. Ciaf1-1, a.k.a. SALK_143656). Among up-regulated proteins were components required for mitochondrial biogenesis, which points to a mitochondrial retrograde signaling pathway being activated and executed in response to the lack of Complex I.