Fig. 5. Localisation of Sla1p and Sla2p in wild-type and mutant cells. (A) KAY303 cells expressing Sla2p and Sla1-myc were grown to log phase and processed for immunofluorescence microscopy as described in Materials and Methods. Both Sla1p-myc-containing and Sla2p-containing patches were polarised to the site of bud emergence, to the enlarging bud and to the bud neck. Furthermore, Sla1p and Sla2p showed substantial co-localisation throughout the cell cycle. However, as indicated by arrows, it was also apparent that a number of cortical patches contained only Sla1p-myc, which suggests that the colocalisation was not always complete. Bar, 10 µM. (B) KAY302 (wildtype) and KAY351 (sla1118-511) cells were grown to log phase and then fixed and processed for immunofluorescence microscopy as described in Materials and Methods. Actin was stained using rhodamine-phalloidin, and Sla2p was localised using antibodies raised to the protein. Bar, 10 µM. (C) The distribution of Sla1-myc in an sla2-null background was examined using immunofluorescence microscopy. In wild-type cells Sla1p was localised to areas of active cell growth, whereas in the absence of sla2 expression Sla1p distribution was depolarised and Sla1p-containing patches appeared evenly distributed around the mother cell and bud. Co-staining with rhodamine-phalloidin revealed that Sla1p-myc was still localised to a subset of cortical patches that contained actin. These images also suggested that in cells lacking Sla2p there are novel structures containing distinct regions of Sla1p and actin organisation (arrows). Bars, 10 µM. The inset shows part of the same image magnified two-fold, and the arrow points toward the same structure in both figures.

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