Fig 8.

Several RNA and protein methyltransferases compete to interact with their cofactor Trm112.(A) Growth plate assay. Serial dilutions (10×) of wild-type cells (BY4741) overexpressing Mtq2, Trm112, or both from pGAL inducible promoters, and an isogenic control, were grown on glucose or galactose-based medium at 30°C for 2 days. +++, overexpression. (B) Western blot analysis of protein steady-state accumulation in strains presented in panel A. Bud23 and Mtq2-Trm112 were detected, respectively, with an antibody raised against the Bud23-Trm112 and Mtq2-Trm112 complexes (see Materials and Methods). Pgk1 (loading control) was detected with a specific commercial antibody. The overexpression of Mtq2 leads to an ∼5-fold reduction in the steady-state accumulation of Bud23 (average calculated on three independent experiments; SD, 0.7). (C) Polysome analysis (254-nm absorbance readings) of strains described for panel A. The arrow points to the 60S peak. The 40S/60S ratio in each panel was determined under dissociation conditions (see Fig. 1C). This experiment was repeated twice. WT, wild type.