AbstractThe conversion of light energy to chemical energy by photosynthesis requires the concerted action of large protein complexes in the thylakoid membrane. Recent work has provided fundamental insights into the three-dimensional structure of these complexes, but how they are assembled from hundreds of parts remains poorly understood. Particularly little is known about the biogenesis of the cytochrome b₆f complex (Cytb₆f), the redox-coupling complex that interconnects the two photosystems. Here we report the identification of a factor that guides the assembly of Cytb₆f in thylakoids of chloroplasts. The protein, DE-ETIOLATION-INDUCED PROTEIN 1 (DEIP1), resides in the thylakoid membrane and is essential for photoautotrophic growth. Knock-out mutants show a specific loss of Cytb₆f, and are defective in complex assembly. We demonstrate that DEIP1 interacts with the two cytochrome subunits of the complex, PetA and PetB, and mediates the assembly of intermediates in Cytb₆f biogenesis. The identification of DEIP1 provides an entry point into the study of the assembly pathway of a crucial complex in photosynthetic electron transfer.