FIGURE 4:

Polysome profiling after loss of Eg5 activity leads to a decrease in the 80S ribosomes and an accumulation of polysomes, indicative of an elongation and/or termination defect. (A) Polysome profiling in RPE1 cells before (left, DMSO + CHX) or after a 4-h 130 μM monastrol treatment (middle, monastrol + CHX) or in the presence of 0.05 mM arsenite (right, arsenite + CHX). CHX was added 10 min prior to cell trypsinization. DMSO + CHX and monastrol + CHX were completed six times, whereas arsenite + CHX was completed twice, all yielding similar results. (B) Same experimental design as in A, except that CHX was omitted. DMSO − CHX and monastrol – CHX were completed four times, whereas arsenite − CHX was completed twice, all yielding similar results. All six of the profiles shown were completed at the same time. Polysome profiling was completed as described in Figure 2A and Materials and Methods. P/M, polysomes/monosomes ratio calculated as described in Materials and Methods.