Sticky Fingers

Connecting Cellular Ca²⁺ With Repolarization

The cardiac action potential is long (even in the atrium) compared to action potentials in other excitable tissues, and the action potential repolarization duration⁴ and configuration⁵ grade cellular Ca²⁺ entry through voltage-gated Ca²⁺ channels (I_Ca). I_Ca is mostly carried by Ca_V1.2 in ventricular myocardium and Ca_V1.2, Ca_V1.3, Ca_V3.1, and Ca_V3.2 in atrium and specialized conduction cells. CaMKII increases I_Ca through CaV1.2,⁶ CaV1.3,⁷ and CaV3.2⁸ and most known intracellular Ca²⁺ homeostatic proteins are CaMKII substrates; therefore, CaMKII is appropriately positioned to serve as a master regulator of cellular Ca²⁺. CaMKII increases I_to to shorten the action potential duration and narrow the membrane potential window permissive for I_Ca. Thus, CaMKII activity at K_V4.3 appears to balance the membrane potential and shorten the action potential to oppose the irrational exuberance of excessive CaMKII-mediated I_Ca increases (Figure, B), at least in healthy myocardium. CaMKII activity supports key physiological tasks in heart, such as excitation–contraction coupling,⁹ and fight or flight automaticity in sinoatrial nodal pacemaker cells.¹⁰ However, excessive CaMKII also contributes to the major diseases of myocardium: arrhythmias and heart failure, leading our group to consider CaMKII as a candidate therapeutic target for heart disease.¹¹

Connecting Cellular Ca²⁺ With Membrane Excitability

CaMKII can modulate membrane excitability by direct effects on Na⁺ current (I_Na) and via indirect effects on the Na⁺/Ca²⁺ exchanger (NCX). I_Na is mostly carried by Na_V1.5 in cardiomyocytes, and CaMKII slows I_Na inactivation to prominently increase a small but important persistent or slow inactivating component of I_Na.¹² CaMKII effects on Na_V1.5 produce a I_Na phenocopy of a rare genetic arrhythmia syndrome (long QT syndrome 3)¹³ and of the increased persistent component of I_Na seen in heart failure.¹⁴ CaMKII activity is increased in heart failure,¹⁵ and an emerging body of evidence supports a view that Na_V1.5 is an important pathological target that contributes to excessive action potential duration, arrhythmia-initiating afterdepolarizations, and subsarcolemmal Na⁺ loading that reduces the efficiency of the NCX for exporting cellular Ca²⁺ to the extracellular space.¹⁶ Whereas I_Na is required for normal membrane excitability in the atrial and ventricular myocardium, NCX current is associated with pathological automaticity in these cells.^17,18 However, the NCX current is now recognized to play an important physiological role in membrane excitability in cardiac pacemaker cells,¹⁹ many of which lack I_Na. Thus, CaMKII is important for augmenting physiological and arrhythmogenic membrane excitability.

Diversity of CaMKII Ion Channel Partnerships

Cellular signaling systems have adapted mechanisms to guide key molecules into close proximity of target proteins. Highly defined localization profiles are an important mechanism for constraining signaling pathways and ensuring specificity. One common theme is for protein kinases to pair with phosphatases by adapter proteins that bring the kinase/phosphatase module to the immediate vicinity of target proteins. We see ultrastructural evidence that CaMKII is targeted to specific microdomains in cardiac myocytes,⁵ and the work of El-Haou et al shows that SAP97 can direct CaMKII signaling in atrial myocytes. In many cases, serine threonine kinases capable of regulating ion channels accomplish localization goals through specialized adapter proteins. In contrast, CaMKII seems not to have committed to a single localization strategy. CaMKII uses diverse approaches, including binding to sequences mimicking the autoinhibitory region of CaMKII itself. The CaMKII autoinhibitory domain is a pseudosubstrate for the catalytic domain and so sequences resembling the autoinhibitory domain have the potential to inhibit²⁰ or anchor²¹ CaMKII by binding the catalytic domain.

The Ca_V1.2 and Ca_V1.3 β subunits are MAGUK²² proteins like SAP97. There are 4 main β subunit genes, and each of these can encode a protein containing a conserved CaMKII phosphorylation site.²³ This CaMKII phosphorylation site (Thr498 on β_2a) is embedded within a primary sequence that is homologous to the CaMKII autoinhibitory domain in β_1b and β_2a. β_1b and β_2a bind CaMKII using these sequences. In fact, the position of Thr498 on β_1b and β_2a mirrors the relationship of the autophosphorylated Thr298 in the CaMKII autoinhibitory domain (Figure, A). CaMKII phosphorylation of Thr498 appears to be necessary for CaMKII effects on Ca_V1.2 pore-forming α subunits.²¹ Interestingly, CaMKII binding to Ca_V1 β subunits is dynamically regulated by Thr498 phosphorylation and by Thr286/287 autophosphorylation. Only autophosphorylated CaMKII binds the Ca_V1 β subunits, whereas phosphorylation of Thr498 on the β subunit markedly disables CaMKII binding.²³ The relationship between Ca_V1.2 activating features of β subunit phosphorylation at Thr498 and the inhibitory consequences of this phosphorylation for CaMKII-β subunit binding suggest a homeostatic mechanism for limiting CaMKII-mediated increases in I_Ca. It will be important for future studies to determine the molecular mechanism for CaMKII binding to K_V channels to better understand the physiological role of CaMKII in modulating K_V channels.

Is CaMKII a Multifunctional β Subunit?

Ion channels exist as macromolecular protein complexes. The α subunits build the channel pore, whereas β subunits act on the pore to influence gating, expression, and assembly of α subunits. We now know that CaMKII is frequently counted among ion channel macromolecular protein complexes, raising the question of whether we should consider CaMKII to be a multifunctional ion channel β subunit. CaMKII performs a wide array of tasks that affect gating, expression, and perhaps intersubunit association.²⁴ Thus, CaMKII is multifunctional based on its repertoire of downstream functions. CaMKII responds to catecholamines²⁰ and angiotensin II² and so provides an important connection between key upstream ligands, membrane excitability, and repolarization. CaMKII has “sticky fingers” that connect a wide range of ion channels to the regulatory influence of Ca²⁺ and reactive oxygen species.