Shr3-NubA interacts with can1-Cub-GST but not hxt1-Cub-GST or gal2-Cub-GST truncations. (A) Schematic diagram of the split ubiquitin truncated constructs used to evaluate Shr3-can1 and Shr3-hxt1/gal2 interactions. (B) Strain FGY135 (gap1Δ shr3Δ) carrying pPL1262 (SHR3-NubA) and pIM46 (can1-8TM-Cub-GST), pIM47 (can1-10TM-Cub-GST), pIM37 (hxt1-8TM-Cub-GST), pIM38 (hxt1-10TM-Cub-GST), pIM43 (gal2-8TM-Cub-GST), or pIM44 (gal2-10TM-Cub-GST) was induced with 2% galactose for 1 h. Extracts were prepared, separated by SGS-PAGE, and analyzed by immunoblotting using antibodies against the 6xHA. (C) The signal intensities of the immunoreactive forms of uncleaved Cub constructs and cleaved interaction marker (GST-6xHA) were quantitated; the mean values of the fraction of split ubiquitin are plotted with error bars showing standard deviation (n = 3; biological replicates). The quantitation corresponding to hxt1-8TM-Cub-GST, hxt1-10TM-Cub-GST, gal2-8TM-Cub-GST, and gal2-10TM-Cub-GST are the same as presented in Fig. 5 C; however, the images correspond to hxt1- and gal2-Cub-constructs are from independent biological replicates. Source data are available for this figure: SourceData F6.