The plasma membrane calcium pump displays memory of past calcium spikes - Differences between isoforms 2b and 4b

To understand how the plasma membrane Ca2+ pump (PMCA) behaves under changi ng Ca2+ concentrations, it is necessary to obtain information about the Ca2 + dependence of the rate constants for calmodulin activation (k(act)) and f or inactivation by calmodulin removal (kin.,at). Here we studied these cons tants for isoforms 2b and 4b. We measured the ATPase activity of these isof orms expressed in Sf9 cells. For both PMCA4b and 2b, k(act) increased with Ca2+ along a sigmoidal curve. At all Ca2+ concentrations, 2b showed a faste r reaction with calmodulin than 4b but a slower off rate. On the basis of t he measured rate constants, we simulated mathematically the behavior of the se pumps upon repetitive changes in Ca2+ concentration and also tested thes e simulations experimentally; PMCA was activated by 500 nm Ca2+ and then ex posed to 50 nm Ca2+ for 10 to 150 s, and then Ca2+ was increased again to 5 00 nm. During the second exposure to 500 nm Ca2+, the activity reached stea dy state faster than during the first exposure at 500 nm Ca2+. This memory effect is longer for PMCA2b than for 4b. In a separate experiment, a calmod ulin-binding peptide from myosin light chain kinase, which has no direct in teraction with the pump, was added during the second exposure to 500 nm Ca2 +. The peptide inhibited the activity of PMCA2b when the exposure to 50 nm Ca2+ was 150 s but had little or no effect when this exposure was only 15 s . This suggests that the memory effect is due to calmodulin remaining bound to the enzyme during the period at low Ca2+. The memory effect observed in PMCA2b and 4b will allow cells expressing either of them to remove Ca2+ mo re quickly in subsequent spikes after an initial activating spike.