• Cytosolic [Ca⁺⁺] is usually less than one micromolar, except during a Ca⁺⁺ signal event.
  Ca⁺⁺-ATPase pumps in the plasma membranes and in endoplasmic reticulum (ER) membranes maintain this low concentration by transporting Ca⁺⁺ away from the cytosol, either out of the cell or into the ER. 
• Extracellular [Ca⁺⁺] in mammalian organisms is in the millimolar range. Opening of plasma membrane Ca⁺⁺ channels may initiate or sustain a Ca⁺⁺ signal.
• [Ca⁺⁺] is also relatively high in the lumen of the ER, which serves as the major internal reservoir from which Ca⁺⁺ is released to the cytosol during Ca⁺⁺ signaling.
• Mitochondria and lysosomes also serve as reservoirs for Ca⁺⁺ subject to release under certain conditions.

Activation of the voltage-gated Ca⁺⁺ channels, by an action potential in the T tubule, leads to opening of ryanodine-sensitive Ca⁺⁺ release channels. Ca⁺⁺ moves from the SR lumen to the cytosol, passing first through the transmembrane portion of the ryanodine receptor, and then through the ryanodine receptor's cytoplasmic assembly. 

• The ryanodine receptor is itself activated by cytosolic Ca⁺⁺ at micromolar concentrations. Thus entry of a small amount of Ca⁺⁺ into the cytosol causes further Ca⁺⁺ release.
• High (e.g., mM) cytosolic Ca⁺⁺ inactivates the ryanodine receptor channel, contributing to signal turn-off.

A somewhat higher resolution structure now available indicates the presence of bent a-helices adjacent to the lumen in the transmembrane pore domain, but an atomic resolution structure of the whole channel has not yet been achieved. For diagrams see article by Ludtke et al. (journal subscription required).
 

The images above were provided by Terrence Wagenknecht of the Wadsworth Center, NY State Dept. of Health. (For references & more information see a Wadsworth Center Web page.)  

In many mammalian cells, IP₃ (inositol-1,4,5-trisphosphate) triggers Ca⁺⁺ release from the endoplasmic reticulum. The "second messenger" IP₃ is produced, e.g., in response to hormonal signals, from the membrane lipid phosphatidylinositol.  (Phosphatidylinositol signals are discussed in more detail elsewhere.)

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Note the stereospecific orientation of substituted and un-substituted hydroxyl groups relative to the two sides of the ring.
Compare to the structure above.

• The IP₃ receptor is a ligand-gated Ca⁺⁺-release channel embedded in endoplasmic reticulum membranes. It is distinct from but partly homologous to the ryanodine receptor channel.
• IP₃ binds to a cytosolic domain of the receptor, promoting channel opening.
  IP₃ may displace a regulatory phospho-protein IRBIT, which binds at the same site.
  See a diagram in a website of the RIKEN Institute, Japan.
• Ca⁺⁺ also binds to the ligand-binding domain of the IP₃ receptor, and promotes channel opening.
  However, high cytosolic Ca⁺⁺, which develops after channel opening, promotes channel closure.
  Thus both the IP₃-activated & ryanodine-sensitive channels are activated by low cytosolic Ca⁺⁺ and inhibited by high cytosolic Ca⁺⁺.
  The feedback inhibition of Ca⁺⁺ release by high cytosolic Ca⁺⁺, along with activity of Ca⁺⁺-ATPase pumps, contributes to signal turnoff and makes possible observed oscillations in Ca⁺⁺ concentration.

Structures of cytosolic domains of the IP₃ receptor, including the IP₃ binding site, have been solved, but the pore structure of the IP₃ receptor has not yet been determined at atomic resolution. 

See image in a website for a low-resolution structure of the IP₃ receptor (AIST, Japan, findings of C. Sato et al.).

of Ca⁺⁺ release &
accumulation by ER 

Calmodulin

Calmodulin, a Ca⁺⁺-activated switch protein, mediates many of the signal functions of Ca⁺⁺. Calmodulin cooperatively binds 4 Ca⁺⁺. 

At each binding site, Ca⁺⁺ interacts with oxygen atoms, mainly of glutamate and aspartate side-chain carboxyl groups, and of the protein backbone, in a loop domain between two a-helices at right angles. This helix-loop-helix motif is called an EF hand (diagram p. 642). 

There are four helix-loop-helix motifs in calmodulin, two at each end of the molecule, which has a dumbbell shape.

Ca⁺⁺ binding to the 4 helix-loop-helix motifs promotes a conformational change that exposes hydrophobic residues along a concave patch on each of the 2 lobes. These residues are involved in protein-protein interactions. Ca⁺⁺-calmodulin then changes conformation again as it wraps around the target domain of a protein.

A typical calmodulin-binding target domain is a positively charged, amphipathic a-helix, with polar and non-polar surfaces. Terminal methyl groups of methionine side-chains of calmodulin participate in binding to hydrophobic residues in target domains of some enzymes that are regulated by calmodulin. However the interaction of Ca⁺⁺-calmodulin with some target proteins is different from what is described here.

Some proteins have bound calmodulin as part of their quaternary structure, even in the absence of Ca⁺⁺. In either case, Ca⁺⁺ binding to calmodulin may induce a conformational change that alters activity of the target protein.

Many enzymes are regulated by Ca⁺⁺-calmodulin. For example:

• Some protein kinases that transfer phosphate from ATP to hydroxyl residues on other enzymes to be regulated are activated by Ca⁺⁺-calmodulin. These are referred to as CaM Kinases.
• The plasma membrane Ca⁺⁺-ATPase that pumps Ca⁺⁺ out of the cell, is one of the target proteins activated by Ca⁺⁺-calmodulin. Thus cytosolic Ca⁺⁺ itself contributes further to turning off Ca⁺⁺ signals.

Additional material on Ca⁺⁺ Signals:
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