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 changing Ca²⁺ concentrations, it is necessary to obtain information about the Ca²⁺ dependence of the rate constants for calmodulin activation (k_act) and for inactivation by calmodulin removal (k_inact). Here we studied these constants for isoforms 2b and 4b. We measured the ATPase activity of these isoforms 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 faster reaction with calmodulin than 4b but a slower off rate. On the basis of the measured rate constants, we simulated mathematically the behavior of these pumps upon repetitive changes in Ca²⁺ concentration and also tested these simulations experimentally; PMCA was activated by 500 nM Ca²⁺ and then exposed to 50 nM Ca²⁺ for 10 to 150 s, and then Ca ²⁺ was increased again to 500 nM. During the second exposure to 500 nM Ca²⁺, the activity reached steady 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 calmodulin-binding peptide from myosin light chain kinase, which has no direct interaction 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 Ca ²⁺ more quickly in subsequent spikes after an initial activating spike.