Functional Analysis Of The MORF Proteins In The Editosome Of Arabidopsis Thaliana Chloroplast

In chloroplasts, primary RNA needs to be modified after transcription, including RNA editing. In recent years, it has been demonstrated that the PPR proteins and some other non-PPR proteins are responsible for RNA editing. MORF family proteins, a class of non-PPR protein which interacts with a lot of PPR proteins, can affect almost all editing sites. RNA editing is previously thought to be carried out by small protein complexes called “editosome”, which including the MORF proteins and PPR proteins. The MORF proteins are the core subunits of the editosome. However, the functional mechanism of the MORF proteins is still unknown. MORF2, MORF8 and MORF9 are found in the chloroplast. In this study, we isolated chloroplast stroma and thylakoid to analyze the location of MORFs. Our results showed that MORF2 mainly localized in chloroplast stroma, while MORF8 and MORF9 mainly exist in thylakoid membrane. Blue-Native results showed that MORF2 and MORF9 exist in the complexes between 67 k Da and 90 k Da, while MORF8 exist in the complexes more than 232 k Da. We screened proteins involved in the editosome by Co-IP, according to the mass spectrometry results. We chose some proteins related to RNA processing to make sure the interactions between those proteins and MORF proteins. The results show that the editosome is not as big as we thought, but MORF2 and MORF9 is close to each other, they may constitute the core of the editosome. Through the RNA immunoprecipitation experiments, we found that the complex of MORF2 and MORF9 are directly involved in the editing of ndh D-2, but MORF2 and MORF9 can not bind RNA directly.