Supplemental Experimental Procedures Strains, Media, and Methods

The S. pombe strains used in this study (Supplemental Table 1.) were grown in YE or minimal medium with appropriate supplements as previously described (Moreno et al., 1991). Strains were constructed by tetrad dissections. DNA transformations were carried out by lithium acetate transformation (Keeney and Boeke, 1994). Induction of the nmt promoter was carried out by initial growth in minimal medium supplemented with thiamine (repressing conditions) and then washing cells three times into minimal medium lacking thiamine (inducing conditions) (Maundrell, 1993). The clp1 ORF was tagged at its 3 ’ end with either the MYC-KanR, HA-KanR, YFP-KanR, or TAP-KanR cassettes as previously described (Bahler et al., 1998). KanR transformants were screened by whole cell PCR and then visualized using fluorescent microscopy or detected through immunoblot analysis to ensure proper integration and expression of the fusion protein. Gene replacements involving the clp1+ ORF were performed as described (Boeke et al., 1984). Briefly, clp1::ura4+ cells were transformed with a genomic construct of clp1 in pIRT2 vector carrying desired mutation(s). Recombinants were selected for in th