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. 2003 Jul;23(13):4449–4460. doi: 10.1128/MCB.23.13.4449-4460.2003

FIG. 5.

FIG. 5.

Nog1 association with pre-60S complexes depends on Rlp24 and is a prerequisite for Nog2 assembly. (A) Early pre-60S complexes accumulate in the absence of Nog1. Rlp24-associated complexes were purified from wild-type (wt) or Nog1-depleted cells (GAL1::NOG1 cells shifted to glucose for 14 h), and the proteins were separated on a 5 to 20% polyacrylamide gel and stained with colloidal Coomassie blue. Asterisks (A to C), positions of the tagged proteins used as baits. The proteins are the same as those in Fig. 2A for comparison. Arrowhead, the Nog2 band identified by mass spectrometry. (B) The role of Nog1 and Rlp24 in Nog2 assembly on pre-60S particles was confirmed by immunoblots with tandem affinity-purified complexes and specific antibodies. Cells were shifted to glucose for 14 h to deplete Nog1 or Rlp24 and for 18 h to deplete Nog2. (C) The requirement of Nog1 for Rlp24 assembly on pre-60S particles was tested by using Ssf1-TAP purified complexes and immunoblots in wild-type cells or after Nog1 or Rlp24 depletion for 14 h.