Abstract
Profilin is an actin- and phosphatidylinositol 4,5-bisphosphate-binding protein that plays a role in the organization of the cytoskeleton and may be involved in growth factor signaling pathways. The subcellular localization of profilin was examined in the yeast Saccharomyces cerevisiae. Immunoblot analysis showed that profilin was localized in both the plasma membrane and cytosolic fractions of the cell. Actin was bound to the profilin localized in the cytosol. The association of profilin with the membrane was peripheral and mediated through interaction with phospholipid. The phospholipid dependence of profilin for membrane binding was examined in vitro using pure profilin and defined unilamellar phospholipid vesicles. The presence of phosphatidylinositol 4,5-bisphosphate in phospholipid vesicles was required for maximum profilin binding. Moreover, the binding of profilin to phospholipid vesicles was dependent on the surface concentration of phosphatidylinositol 4,5-bisphosphate. The subcellular localization of profilin was examined in vivo under growth conditions (i.e. inositol starvation of ino1 cells and glucose starvation of respiratory deficient cells) where plasma membrane levels of phosphatidylinositol 4,5-bisphosphate were depleted. Depletion of plasma membrane phosphatidylinositol 4,5-bisphosphate levels resulted in a translocation of profilin from the plasma membrane to the cytosolic fraction. Profilin translocated back to the membrane fraction from the cytosol under growth conditions where plasma membrane levels of phosphatidylinositol 4,5-bisphosphate were replenished. These results suggested that phosphoinositide metabolism played a role in the localization of profilin.
Highlights
Profilin is an actin- and phosphatidylinositol 4,5bisphosphate-binding protein that plays a role in the organization of the cytoskeleton and may be involved in growth factor signaling pathways
About 80% of the total amount of profilin was associated with membranes, whereas the remainder was associated with the cytosolic fraction (Fig. 1B)
In this work we examined the regulation of profilin localization by PIP2 metabolism in S. cerevisiae
Summary
Profilin is an actin- and phosphatidylinositol 4,5bisphosphate-binding protein that plays a role in the organization of the cytoskeleton and may be involved in growth factor signaling pathways. Profilin translocated back to the membrane fraction from the cytosol under growth conditions where plasma membrane levels of phosphatidylinositol 4,5-bisphosphate were replenished These results suggested that phosphoinositide metabolism played a role in the localization of profilin. Profilin is required for the proper organization of the actin cytoskeleton into actin cables, which occur at regions of active growth and for proper maintenance of cell polarity [12] It is unclear whether these roles for profilin are related to its ability to bind actin and/or PIP2 [13]. The results of our studies were consistent with the notion that the metabolism of PIP2 played a role in the localization of profilin
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