A Ycf2-FtsHi heteromeric AAA-ATPase complex is required for chloroplast protein import

Chloroplasts import thousands of nucleus-encoded preproteins synthesized in the cytosol through the TOC and TIC translocons on the outer and inner envelope membranes, respectively. Preprotein translocation across the inner membrane requires ATP; however, the import motor has remained unclear. Here, we report that a 2-MDa heteromeric AAA-ATPase complex associates with the TIC complex and functions as the import motor, directly interacting with various translocating preproteins. This 2-MDa complex consists of a protein encoded by the previously enigmatic chloroplast gene Ycf2 and five related nuclear-encoded FtsH-like proteins, namely FtsHi1, FtsHi2, FtsHi4, FtsHi5, and FtsH12. These components are each essential for plant viability and retain the AAA-type ATPase domain, but only FtsH12 contains the zinc-binding active site generally conserved among FtsH-type metalloproteases. Further, even the FtsH12 zinc-binding site is dispensable for its essential function. Phylogenetic analyses suggest that all AAA-type members of the Ycf2/FtsHi complex including Ycf2 evolved from the chloroplast-encoded membrane-bound AAA-protease FtsH of the ancestral endosymbiont. The Ycf2/FtsHi complex also contains an NAD-malate dehydrogenase, a proposed key enzyme for ATP production in chloroplasts in darkness or in non-photosynthetic plastids. These findings advance our understanding of this ATP-driven protein translocation system that is unique to the green lineage of photosynthetic eukaryotes.