Immunoblot with anti-Sds22 antiserum of yeast extracts from (1) wild type strain HM123, (2) sds::ura4+ deletion mutant carrying pHR140-2. The 40kD protein band was identified by immunoblot analysis of wild-type strain using anti-Sds22 antisera (lane1). The 40 kD band is enhanced in the sds22::ura4+ disruptrion mutant strain that is rescued by the multicopy sds22+ plasmid pHR140-2 (lane2).

Sds22 subcellular localization Indirect immunofluorescence microscopy was performed by staining methanol fixed cells with (first column) anti-Sds22 antiserum, or (second column) DAPI to visualize chromosomal DNA. (a) wild type HM123; (b) HM123 carrying multicopy sds22+ plasmid pHR140-2. Anti-Sds22 antibody stains the cytoplasm as well as the non-chromosomal domain of the nucleus of a wild type strain, as shown in (a). Nuclear staining increases in strains carrying a multicopy sds22+ plasmid (b).

Sds22 coprecipitates with Dis2 and Sds21. Yeast extracts of wild type (wt) strain HM123 (lane 1 and 2), dis2::ura4+ deletion mutant (Δd2, lane 3), sds21::ura4+ deletion mutant (Δs21, lane 4) were immunoprecipitated followed by immunoblotting with the indicated antiserum, to detect the Sds22 or Dis2/Sds21 proteins. Lane 1 was immunoprecipitated with the appropriate preimmune serum, lane 2-4 with the anti-Sds22 serum. (a) denotes anti-Sds22 immunoblot; (b) denotes anti-D2C immunoblot. Anti-D2C crossreacts with both Sds21 and Dis2. Anti-Sds22 antiserum coprecipitates both Dis2 and Sds21 proteins in the wild type strain (lane 2b). Consistently, Sds21 alone is precipitated in the dis2 deletion mutant (lane 3b), and Dis2 alone is precipitated in the sds21 deletion mutant (lane 4b).