SPR2 interacts with multiple splicing factors. A, Interaction analysis of between SPR2 and EMP16, PPR14, and Zm-mCSF1 in Y2H assays. DDO medium: SD/–Leu/–Trp medium; QDO medium: SD/–Ade/–His/–Leu/–Trp medium; QDO/X/A: QDO medium with added the X-α-gal and AbA. AD: GAL4 activation domain; BD: GAL4 DNA binding domain. B, LCI assays to determine interactions between SPR2 and EMP16, PPR14, and Zm-mCSF1. The intensity of the fluorescent signals represents their interaction activities. C, BiFC analysis for interactions between SPR2 and EMP16, PPR14, and Zm-mCSF1 in N. benthamiana leaves. YFP signals were detected by a confocal laser microscope. Scale bars, 20 μm. D, A proposed working model of SPR2. Different colors represent various splicing factor. PPRs indicates other P-type PPR proteins, CRMs indicates CRM family proteins, and question mark indicates the other kind of mitochondrial splicing factors.