FIGURE 10.
PhoClp1 can perform the 5′ end healing step of tRNA splicing in yeast. Yeast trl1Δ p360–TRL1 (URA3 CEN) cells were cotransformed with a CEN TRP1 plasmid bearing AtRNL–S701A plus either: (1) p[CEN HIS3 TRL1-(389–827)] expressing the kinase–CPD domain of yeast tRNA ligase (positive control), (2) an empty 2μ HIS3 vector (negative control), or (3) p[2μ HIS3 PhoCLP1] expressing PhoClp1 under the control of the yeast TPI1 promoter. Trp+ His+ transformants were selected at 30°C and then streaked on agar medium containing 0.75 mg/mL 5-fluoroorotic acid. The plates were photographed after incubation for 4 d at 30°C.
