Voltage-gated Ca2+ Channels (VGCCs)

For review, see:

Catterall, 2000. Annu. Rev. Cell Dev. Biol. 16: 521-555.

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• Ca2+-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 Ca2+/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., 2004. Neuron 42:89-99

Introduction• The subunits are cytosolic components of

Ca2+ 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 Ca2+ channels

Kim et al., 2004. Neuron 41: 745-754.

Introduction• Ca2+ entry is limited by Ca2+-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 Ca2+/CaM effector site.

• EF hand, classically thought of as a Ca2+ binding site.

• 110 amino acids in between IQ and EF:

• Peptide A = 1588-1609; can bind CaM in absence of Ca2+

• Peptide C = binds CaM with k1/2 for Ca2+ < 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 = IBa

Gray line = ICa

WT = - - - - Mutant =

Figure 4 - Kim

Black line = IBa

Gray line = ICa

WT = - - - - Mutant =

ApoCaM tethering is not necessary nor sufficient for producing accerated VDI.

Figure 5 - Kim

Figure 6 - Kimm

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tw

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Conclusions – Kim

• C terminal apoCaM tethering domains and Ca2+/CaM effector domains that regulated CDI are inseparable.

Control of ion conduction in L-type Ca2+ channels by the

concerted action of S5-6 regions

Cibulsky and Sather, 2003. Biophys J. 84: 1709-1719.

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 mV1s wt = 67.7 mV

sQuadS5-6c = 46.3 mVcQuadPs = 61.1 mVcQuadS5-6s = 63.9 mV

Fig. 2: Cd2+ Block

Fig. 3: P loop transfer from 1S to 1C

1C wt = 28.9 pS 1s wt = 16.3 pScQuadPs = 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

cQuadS5-6s = 14.1 pS

Fig.5: Reciprocal transfer - 1C to 1S

1C wt = 28.9 pS

1s wt = 16.3 pS

sQuadS5-6c = 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 Ca2+ 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.