Cytochrome b (Cyt b) is an essential component of the photosystem II (PSII) reaction center core. It consists of two subunits, PsbE and PsbF, which together coordinate a redox-active heme. While extensive studies have revealed the importance of Cyt b, its structural and functional roles are not fully understood. Previous studies have implied that the lumenal region of Cyt b, interacting with the PSII extrinsic subunit PsbP in green plant PSII, may have important roles. However, few studies have investigated its lumenal region. Here, we have focused on a well-conserved lumenal region of PsbE, which was found to interact with the N-terminal region of PsbP in green-lineage PSII (from green algae and land plants). In red-lineage PSII (from red algae and algae possessing red algal-derived plastids), very similar interactions were observed between the same lumenal region of PsbE and the N-terminal region of PsbQ'. We generated Arabidopsis thaliana mutants harboring mutations in the well-conserved lumenal region of PsbE through targeted base editing of the plastid genome by ptpTALECD. The mutations led to strong growth defects and extremely low F/F. This study suggests the importance of the lumenal regions of Cyt b, and gives insight into possible structural and functional compensation between the N-terminal regions of PsbP in green-lineage PSII and PsbQ' in red-lineage PSII.