Conserved Kv4 N-terminal Domain Critical for Effects of Kv Channel-interacting Protein 2.2 on Channel Expression and Gating

Abstract

Nature 403, 553-556). We cloned a KChIP2 splice variant (KChIP2.2) from human ventricle. In comparison with KChIP2.1, coexpression of KChIP2.2 with human Kv4 channels in mammalian cells slowed the onset of Kv4 current inactivation (2-3-fold), accelerated the recovery from inactivation (5-7-fold), and shifted Kv4 steady-state inactivation curves by 8-29 mV to more positive potentials. The features of Kv4.2/KChIP2.2 currents closely resemble those of cardiac rapidly inactivating transient outward currents. KChIP2.2 stimulated the Kv4 current density in Chinese hamster ovary cells by ϳ55-fold. This correlated with a redistribution of immunoreactivity from perinuclear areas to the plasma membrane. Increased Kv4 cell-surface expression and current density were also obtained in the absence of KChIP2.2 when the highly conserved proximal Kv4 N terminus was deleted. The same domain is required for association of KChIP2.2 with Kv4 ␣-subunits. We propose that an efficient transport of Kv4 channels to the cell surface depends on KChIP binding to the Kv4 N-terminal domain. Our data suggest that the binding is necessary, but not sufficient, for the functional activity of KChIPs.Voltage-gated potassium (Kv) channels related to the Shal (Kv4) gene family (2-5) mediate rapidly inactivating outward currents related to neuronal subthreshold A-type currents (4) as well as transient outward currents (I to ) in cardiac myocytes (6). Kv4 channel subunits have been localized immunocytochemically to somatodendritic areas in rat brain neurons (7). The high abundance of neuronal Kv4 channels in neuronal soma and dendrites suggests that Kv4 channels play an important role in postsynaptic signal integration. In fact, Kv4 channel activity may prevent action potential back-propagation into dendrites, thereby controlling potentiation of dendritic excitability (8) and acting as "dendritic shock-absorbers" (9). Conversely, inactivation of Kv4 channels below the action potential firing threshold leads to increased postsynaptic neuronal excitability (10, 11).Recently, small Ca 2ϩ -binding and Kv channel-interacting proteins (KChIPs) 1 were discovered (1). They are encoded in three different genes (KChIP1, KChIP2, and KChIP3) (1) and are members of the recoverin superfamily of Ca 2ϩ -binding proteins (12), being closely related to frequenin (13). KChIPs are associated with Kv4 channels as shown by immunocytochemical colocalization and co-immunoprecipitation studies (1). Since KChIPs are prominently expressed in brain, neuronal Kv4 channels most likely contain KChIPs as an integral subunit component.Kv4 channels are also expressed in cardiac tissue, where they mediate I to . This current contributes to the early phase of cardiac action potential repolarization (14). We considered the possibility that, like neuronal Kv4 channels, cardiac Kv4 channels might be associated with KChIPs. Accordingly, we searched human cardiac mRNA for KChIP mRNA. In this work, we report the cloning of a human cardiac KChIP cDNA, KChIP2.2, which represents a splice variant...

Kv channel-interacting proteins as neuronal and non-neuronal calcium sensors

“…An and coworkers [1] showed that the KChIP1ΔEF2,3,4 mutant was unable to modulate the inactivation gating of Kv4.2, however, these authors reported (without showing the data) that KChIP1ΔEF2,3,4 coimmunoprecipitated with the Kv4.2 α-subunit, supporting the notion that Kv4.2/KChIP1 complex formation is not Ca 2+ -dependent. Apparently, this applies also to Kv4.2/KChIP2 complex formation, because Bähring and coworkers [70] found that binding of KChIP2c to a 180 amino acid NTF of Kv4.2 occurs both in the absence and presence of Ca 2+ . Structural differences between Kv4.3 and Kv4.2, differences in KChIP binding efficiency between NTFs and Kv4 full-length protein, and, not least, structural differences between different KChIP subtypes, may be responsible for the seemingly contradictory results.…”

“…(E) Graphs show the effects of Kv4/KChIP coexpression (black: Kv4 alone, grey: Kv4 + KChIP). Shown data (increase in current density, acceleration of initial curent decay kinetics, acceleration of recovery from inactivation and positive shift of the voltage dependence of steady-state inactivation) refer to Kv4.2 + KChIP2c coexpression [70]. Red arrows indicate that Ca 2+ binding to KChIPs may have an influence on the observed effects.…”

“…Heterologous coexpression of KChIPs (except for the KChIP4a splice variant, see below) leads to an increase in surface expression of Kv4 channels and to a modulation of their inactivation gating, namely, a slowing of the initial phase of macroscopic current decay, an acceleration of the recovery from inactivation and a positive shift of the voltage dependence of steady-state inactivation (Figure 1(e)). [1,70,71] As for any other type of NCS protein and its target molecule(s), a putative calcium sensor function of KChIPs related to Kv4 channels [1] may be mediated in various, not mutually exclusive, ways: One possibility is that Kv4/KChIP complex formation itself depends on the Ca 2+ binding state of the KChIP β-subunit. This aspect also includes the intriguing possibility of a Ca 2+ -dependent dynamic Kv4:KChIP stoichiometry.…”

“…It has been recognized early on that, with the exception of the KChIP4a splice variant, complex formation with KChIPs promotes the trafficking of Kv4 channels to the plasma membrane and increases their stability [1,70,79]. It was suspected that some ER retention motif within the Kv4 N-terminus is masked, and thus rendered ineffective, during Kv4/KChIP complex formation [70,79].…”

“…It was suspected that some ER retention motif within the Kv4 N-terminus is masked, and thus rendered ineffective, during Kv4/KChIP complex formation [70,79]. In such a scenario Ca 2+ -sensitive KChIP binding to the Kv4 N-terminus (see above) would also confer Ca 2+ sensitivity to channel trafficking.…”