Voltage-gated Ca 2+ Channels (VGCCs) For review, see: Catterall, Annu. Rev. Cell Dev. Biol. 16:

Voltage-gated calcium channels Calcium is “THE ION” because of its physiological role in nearly every cellular process, including: Gene regulation Signal transduction Neurotransmitter release Hormone secretion Ca 2+ -dependent action potentials Fertilization Cell death (apoptosis) Modulation of ion channel activity Excitation-contraction coupling (muscle) and on and on and on…

Voltage-gated calcium channels These channels are regulated by: Phosphorylation (cAMP-dependent protein kinase) G proteins (uncommon modulation by G  Calcium and Ca 2+ /CaM Intracellular effector proteins (such as the RyR, SNARE proteins)

Voltage-gated calcium channels

Figure 3 - Catterall

Calcium channel function regulated by the SH3-GK module in  subunits McGee et al., Neuron 42:89-99

Introduction The  subunits are cytosolic components of Ca 2+ channels that are necessary for proper expression and kinetics of the  subunit. There are two conserved regions of the  subunit: –C1 –C2 –Three variable regions (V1-3) flank C1/C2 and are targets for postranslational modification (e.g., phosphorylation/palmitoylation)

Figure 1 - McGee

Figure 2 - McGee

Figure 3 - McGee

Figure 4 - McGee

Table 1 - McGee

Figure 5 - McGee

Figure 6 - McGee

Figure 7 - McGee

Conclusions – McGee  subunits are similar to MAGUKs; they contain a split SH3 fold that can assemble from subdomains composed to C1 (  -SH3) and C3 (  -GK) regions in either an intra- or intermolecular fashion.

Identification of the components controlling inactivation of voltage-gated Ca 2+ channels Kim et al., Neuron 41:

Introduction Ca 2+ entry is limited by Ca 2+ -dependent inactivation (CDI). CDI depends on constitutively bound calmodulin (CaM). apoCaM = calmodulin lacking bound calcium Question: How do CaM and the channel form a calcium-sensing apparatus???

Figure 1 - Kim IQ motif. In C terminus of pore-forming  subunit. Acts as a Ca 2+ /CaM effector site. EF hand, classically thought of as a Ca 2+ binding site. 110 amino acids in between IQ and EF: Peptide A = ; can bind CaM in absence of Ca 2+ Peptide C = binds CaM with k 1/2 for Ca 2+ < 90 nM

Introduction Question: do calcium-dependent inactivation (CDI) and voltage-dependent inactivation (VDI) utilize the same machinery, a cytoplasmic I-II linker, to form a blocking peptide?

Figure 2 - Kim

Figure 3 - Kim

Figure 4 - Kim Black line = I Ba Gray line = I Ca WT = Mutant =

Figure 4 - Kim Black line = I Ba Gray line = I Ca WT = Mutant = ApoCaM tethering is not necessary nor sufficient for producing accerated VDI.

Figure 5 - Kim

Figure 6 - Kim mutant wt

Conclusions – Kim C terminal apoCaM tethering domains and Ca 2+ /CaM effector domains that regulated CDI are inseparable.

Control of ion conduction in L-type Ca 2+ channels by the concerted action of S5-6 regions Cibulsky and Sather, Biophys J. 84:

Figure 1 - Cibulsky

Fig. 2: Activation  1C : fast activation (as expected)  1S : slow activation (as expected)  1S based, No change  1C based, No change

Fig. 2: Reversal Potential Reversal  :  1C wt = 73 mV  1s wt = 67.7 mV s QuadS5-6 c = 46.3 mV c QuadP s = 61.1 mV c QuadS5-6 s = 63.9 mV

Fig. 2: Cd 2+ Block

Fig. 3: P loop transfer from  1S to  1C  1C wt = 28.9 pS  1s wt = 16.3 pS c QuadP s = 22.9 pS Conclusion: additional parts of the channel affect unitary conductance.

Fig. 4: S5-6 transfer from  1S to  1C  1C wt = 28.9 pS  1s wt = 16.3 pS c QuadS5-6 s = 14.1 pS

Fig.5: Reciprocal transfer -  1C to  1S  1C wt = 28.9 pS  1s wt = 16.3 pS s QuadS5-6 c = 30.0 pS

Fig. 6

Conclusions - Cibulsky S5-6 region contains the structural features that are responsible for the difference in unitary conductance between  1S and  1C L-type Ca 2+ channels. The pore region alone does not confer all properties of unitary conductance. Reciprocal swap indicates that no other regions account for the characteristic ion transport rates of the two types of channels.