Plasma membrane calcium pump activity is affected by the membrane protein concentration: Evidence for the involvement of the actin cytoskeleton

Plasma membrane calcium pumps (PMCAs) are integral membrane proteins that actively expel Ca2+ from the cell. Specific Ca2+- ATPase activity of erythrocyte membranes increased steeply up to 1.5-5 times when the membrane protein concentration decreased from 50 mu g/ml to 1 mu g/ml. The activation by dilution was also observed for ATP-dependent Ca2+ uptake into vesicles from Sf9 cells overexpressing the PMCA 4b isoform, confirming that it is a property of the PMCA. Dilution of the protein did not modify the activation by ATP, Ca2+ or Ca2+-calmodulin. Treatment with non-ionic detergents did not abolish the dilution effect, suggesting that it was not due to resealing of the membrane vesicles. Pre-incubation of erythrocyte membranes with Cytochalasin D under conditions that promote actin polymerization abolished the dilution effect. Highly-purified, micellar PMCA showed no dilution effect and was not affected by Cytochalasin D. Taken together, these results suggest that the concentration-dependent behavior of the PMCA activity was due to interactions with cytoskeletal proteins. The dilution effect was also observed with different PMCA isoforms, indicating that this is a general phenomenon for all PMCAs. (c) 2007 Elsevier B.V. All rights reserved.