Figure 5. Not5 co-precipitates with Csl4 and Rrp41.

A. Total protein extracts were prepared from wild-type or caf40Δ cells expressing Tap-tagged Csl4. 2 mg of total protein was incubated with (IP) or without (IP₀) antibodies against CBP. 50 µg of total extract (TE), equivalent volume of unbound extract (FT) and the immunoprecipitate were loaded on SDS-PAGE followed by western blotting with antibodies against Not5, which revealed both Not5 and Tap-tagged Csl4 as indicated. B. 2 g of total protein extract from wild-type cells expressing Tap-tagged Rrp41 treated or not with RNAse as indicated were processed for tandem affinity purification of Rrp41. Equivalent amounts of total extract (TE), flow through (FT) or one fourth of the eluate precipitated by TCA, were loaded on SDS-PAGE and analyzed with antibodies against CBP to follow Rrp41, or against Rrp4, Rrp43 and Not5 as indicated.