FIGURE 4.

Rdn1 and Rdn2 assemble as distinct RDRCs that share association with Dcr2. (A) Strains expressing tagged Rdn1 or Rdn2 were created using two strategies: introduction of a C-terminal tag cassette at the endogenous locus or integration of an N-terminally tagged protein transgene at the nonessential BTU1 locus. Neo-S and Neo-R or Taxol-S and Taxol-R indicate relative phenotypic sensitivity and resistance to the selective drug. (B) Protein complexes were examined following purification of the indicated tagged proteins from extracts of starved cells (lanes 1,4–5) or cells at the 9-h time point of conjugation (lanes 2,3). Parallel mock purifications were performed using wild-type cell extracts, only one of which is shown but all of which were similar in nonspecific background. The composition of the multisubunit complexes identified here is summarized on the right. Lighter text denotes the RDRC subunits that exhibit increased expression during conjugation. (C) Dcr2 association with Rdr1 was examined following purification of the indicated tagged proteins from extracts of starved cells using the gentle washing conditions that preserve RDRC-Dcr2 interaction.