The apicoplast of Plasmodium parasites serves as a metabolic hub that synthesize essential biomolecules. Like other endosymbiotic organelles, 90% of the apicoplast proteome is encoded by the cell nucleus and transported to the organelle. Evidence suggests that the apicoplast has minimal control over the synthesis of its proteome and therefore it is unclear how organelle proteostasis is regulated. Here, we identified and investigated a large and conserved chaperonin (CPN) complex with a previously unknown function. Using genetic tools, we demonstrated that its main subunit, CPN60, localizes to the apicoplast and is essential for parasite's asexual replication due to its organellar functions. Unlike its close orthologues in other prokaryotic and eukaryotic cells, CPN60 is not upregulated during heat shock (HS) and does not affect HS response in the parasite. Instead, we found that it is directly involved in proteostasis through interaction with a proteolytic complex called Clp (caseinolytic protease) that degrades organellar proteins in a regulated manner. We showed that CPN60 physically binds both the active and inactive forms of the Clp complex, and manipulates its stability. Using computational prediction tools, we modeled the interaction between these two large complexes to understand how a stable interface is attained. Finally, we screened a library of inhibitors for the bacterial CPN60 orthologue GroEL, in order to test the potential of chaperonin inhibition as antimalarial. These inhibitors demonstrated an anti-Plasmodial activity that was not restricted to apicoplast function. Taken together, this work reveals how balanced activities of proteolysis and refolding safeguard the apicoplast proteome.
