Figure 3.

J domain and G/F region of Drosophila and human Sis1 homologs can complement Sis1's roles in cell growth and [RNQ⁺] maintenance. (A) A 10-fold dilution series of cell suspensions from a Δsis1 (left) or ydj1 (right) strain carrying SIS1 or YDJ1, respectively, on a URA3-containing vector and second plasmid having no Hsp40 gene (−), SIS1, Drosophila DROJ1, human HDJ1, or human HDJ2. Cells were spotted on media containing 5-FOA and incubated for 3 d. (B–D) Top, fluorescence microscopy of Δsis1 cells coexpressing an Hsp40 gene and RNQ1-GFP. Cells were visualized 4 h after RNQ1-GFP induction as described above. Bottom, aggregation assays for the lysates of corresponding strains. After centrifugation, equivalent aliquots of total lysate T, supernatant fraction S, and pellet fraction P were subjected to immunoblot analysis by using Rnq1-specific antibody. (B) Full-length DROJ1 and HDJ1. (C) Truncation mutants of DROJ1 and HDJ1, droJ1-143 and hdj1-147, which encompass their corresponding J domain and glycine-rich sequences. (D) Sis1-121 truncation or a fusion that includes the J domain of Ydj1 fused to the G/F region of Sis1 (YS-121).