TY - JOUR
T1 - A bimodular mechanism of calcium control in eukaryotes
AU - Tidow, Henning
AU - Poulsen, Lisbeth Rosager
AU - Andreeva, Antonina
AU - Knudsen, Michael
AU - Hein, Kim Langmach
AU - Wiuf, Carsten Henrik
AU - Palmgren, Michael Broberg
AU - Nissen, Poul
PY - 2012/11/15
Y1 - 2012/11/15
N2 - Calcium ions (Ca 2+) have an important role as secondary messengers in numerous signal transduction processes, and cells invest much energy in controlling and maintaining a steep gradient between intracellular (∼0.1-micromolar) and extracellular (∼2-millimolar) Ca 2+ concentrations. Calmodulin-stimulated calcium pumps, which include the plasma-membrane Ca 2+ -ATPases (PMCAs), are key regulators of intracellular Ca 2+ in eukaryotes. They contain a unique amino- or carboxy-terminal regulatory domain responsible for autoinhibition, and binding of calcium-loaded calmodulin to this domain releases autoinhibition and activates the pump. However, the structural basis for the activation mechanism is unknown and a key remaining question is how calmodulin-mediated PMCA regulation can cover both basal Ca 2+ levels in the nanomolar range as well as micromolar-range Ca 2+ transients generated by cell stimulation. Here we present an integrated study combining the determination of the high-resolution crystal structure of a PMCA regulatory-domain/calmodulin complex with in vivo characterization and biochemical, biophysical and bioinformatics data that provide mechanistic insights into a two-step PMCA activation mechanism mediated by calcium-loaded calmodulin. The structure shows the entire PMCA regulatory domain and reveals an unexpected 2:1 stoichiometry with two calcium-loaded calmodulin molecules binding to different sites on a long helix. A multifaceted characterization of the role of both sites leads to a general structural model for calmodulin-mediated regulation of PMCAs that allows stringent, highly responsive control of intracellular calcium in eukaryotes, making it possible to maintain a stable, basal level at a threshold Ca 2+ concentration, where steep activation occurs.
AB - Calcium ions (Ca 2+) have an important role as secondary messengers in numerous signal transduction processes, and cells invest much energy in controlling and maintaining a steep gradient between intracellular (∼0.1-micromolar) and extracellular (∼2-millimolar) Ca 2+ concentrations. Calmodulin-stimulated calcium pumps, which include the plasma-membrane Ca 2+ -ATPases (PMCAs), are key regulators of intracellular Ca 2+ in eukaryotes. They contain a unique amino- or carboxy-terminal regulatory domain responsible for autoinhibition, and binding of calcium-loaded calmodulin to this domain releases autoinhibition and activates the pump. However, the structural basis for the activation mechanism is unknown and a key remaining question is how calmodulin-mediated PMCA regulation can cover both basal Ca 2+ levels in the nanomolar range as well as micromolar-range Ca 2+ transients generated by cell stimulation. Here we present an integrated study combining the determination of the high-resolution crystal structure of a PMCA regulatory-domain/calmodulin complex with in vivo characterization and biochemical, biophysical and bioinformatics data that provide mechanistic insights into a two-step PMCA activation mechanism mediated by calcium-loaded calmodulin. The structure shows the entire PMCA regulatory domain and reveals an unexpected 2:1 stoichiometry with two calcium-loaded calmodulin molecules binding to different sites on a long helix. A multifaceted characterization of the role of both sites leads to a general structural model for calmodulin-mediated regulation of PMCAs that allows stringent, highly responsive control of intracellular calcium in eukaryotes, making it possible to maintain a stable, basal level at a threshold Ca 2+ concentration, where steep activation occurs.
U2 - 10.1038/nature11539
DO - 10.1038/nature11539
M3 - Letter
C2 - 23086147
SN - 0028-0836
VL - 491
SP - 468
EP - 472
JO - Nature
JF - Nature
IS - 7424
ER -