FIGURE 9:

Cbk1 inhibition prolongs Bck2 and Sdp1 puncta localization during heat shock. (A) Bck2-GFP localization was monitored at various intervals after heat shock in wild-type cells (left) and in cbk1-as cells upon Cbk1 inhibition in cbk1-as cells (right). For cbk1-as cells, Cbk1 inhibition (1NA-PP1 addition) and heat shock were done simultaneously. NLS-RFP shows the nuclear localization. Bck2 localized to cytoplasmic puncta in all cells (n = 35) within 15 min of heat shock. All images were captured via wide-field fluorescence microscopy and represent two merged optical sections. See Supplemental Figure S3 for a temporal analysis of Bck2 localization at mitochondria. (B) Parallel experiments were done in cells coexpressing Sdp1-RFP, and the number of cells with three or more Bck2 and Sdp1 cytoplasmic puncta was plotted over time. The data were tabulated from two independent experiments (n = 29–55 cells per time point). Cbk1 kinase inhibition significantly delayed the disappearance of the heat shock–induced Bck2 and Sdp1 puncta. See Supplemental Figure S4 for additional controls. The strains used for these experiments are FLY3503 and FLY3559.