Figure 9.

OZ2 interacts with several site-specific and general mitochondrial splicing factors in vitro and in vivo. (A) Yeast two-hybrid assay for OZ2/mitochondrial splice factors demonstrate interactions with certain factors that share splicing targets with OZ2, as well as OZ2 dimerization. Haploid yeast were transformed with constructs expressing OZ2 or a mitochondrial splicing factor with N-terminal fusions of parts of the yeast GAL4 transcription factor. Transformed yeast were mated to produce diploids expressing two fusion proteins and selected on synthetic dropout media lacking leucine and tryptophan (-L-W). Interactions were tested on plates lacking leucine, tryptophan, histidine, and adenine (-L-W-H-Ade). Yeast dilution spots contained 10⁶, 10⁵ and 10⁴ cells per mL. Em = empty vector. (B-I) Bimolecular fluorescence complementation (BiFC) was performed in N. benthamiana using transient nYFP/cYFP fusion protein expression constructs. Green = YFP fluorescence; magenta = chlorophyll autofluorescence; scale bars = 10 μm. (B) ABO5 + OZ2. (C) BIR6 + OZ2. (D) MAT2 + OZ2. (E) mCSF1 + OZ2. (F) MISF26 + OZ2. (G) MTL1 + OZ2. (H) PMH2 + OZ2. (I) OZ2 + OZ2. The interaction of OZ2 with itself serves as a positive control for this assay.