KChIPs (Kv channel-interacting proteins) — a few surprises and another

Excitable cells vary in the pattern of electrical activity they spontaneously produce. Voltage-gated K⁺ currents are very diverse and are often considered to be major determinants of excitability. The presence of a large number of Kv channel subunit genes suggests that electrophysiological phenotype is primarily determined by the assortment of channel genes expressed by each cell. While this notion is still true in general, the last decade has seen the addition of a few more layers of complexity in generating diverse Kv channels. One of the most important additions is the identification of various auxiliary and other channel-interacting proteins. In 2000, a study of the channels in the Kv4 family added new members to this ever-increasing list of channel auxiliary subunits (An et al. 2000). These Kv4 channel-interacting proteins are named KChIPs. KChIPs are encoded by at least four genes, KChIP1-4: all four KChIP genes are highly expressed in the brain, whereas only KChIP2 mRNA is abundant in the heart. Association with KChIPs dramatically increases current amplitude and alters the gating properties of Kv4 channels. Kv4-KChIP complexes exhibit some of the gating properties and modulation which are reminiscent of native transient Kv currents in cardiac myocytes and neurons. Furthermore, differential expression of the KChIP2 gene is well correlated with the density of the transient outward K⁺ current across the left ventricular wall of large animals. Deletion of the KChIP2 gene leads to the loss of this current. These findings indicate that KChIPs are essential components of the channels carrying the transient outward K⁺ current in cardiac myocytes and somatodentritic A-type current in some neurons.

While the studies on KChIPs have shown that they are physiologically important auxiliary subunits of Kv4 channels, several surprises have been revealed. The first unexpected finding was the fact that KChIPs are Ca²⁺-binding proteins. They belong to a family of four EF-hand motif-containing proteins including recoverin, guanylyl cyclase-activating proteins (GCAP), neurocalcins or visinin-like proteins (VILIPs), and neuronal calcium sensor-1 (NCS-1) corresponding to yeast frequenin. However, it is not generally thought that Ca²⁺ modulates the transient currents in cardiac myocytes or neurons, although most of the current measurements are done in the presence of a Ca²⁺ chelator in order to eliminate the contamination of Ca²⁺-activated currents. A further surprise came from two completely unrelated studies: in addition to acting as an auxiliary subunit for Kv channels, KChIP3 has been shown to function as a transcription factor (Carrion et al. 1999) and a regulator of the Alzheimer's disease-associated proteins presenilins (Buxbaum et al. 1998). There are some worries associated with these studies. For example, the proposed DNA cis-element seems to have low sequence specificity and affinity for the binding to KChIP3. Likewise, the in vivo presence of presenilin-KChIP3 complexes has never been demonstrated. Despite these uncertainties, the findings raise an intriguing possibility that KChIP3, as well as its related Ca²⁺-binding proteins, may control multiple cellular functions. Finally, recent studies have indicated that NCS-1 produces similar changes in Kv4 channel gating to KChIPs (Nakamura et al. 2001; Guo et al. 2002). Anti-NCS-1 antibody appeared to coimmunoprecipitate Kv4 channel proteins from neonatal heart extract (Guo et al. 2002). These findings are unexpected, because An et al. (2000) using yeast two hybrid assays, found that the N-terminal peptide of Kv4.2 protein specifically interacts with KChIPs, but not NCS-1. Nevertheless, these studies indicate that not only KChIPs, but also NCS-1, may be physiological regulators of Kv4 channels in native cells.

Another surprise comes in this issue of The Journal of Physiology. Patel et al. (2002) report the identification of a truncated form of KChIP2 (KChIP2d) that consists of only the latter half of the previously known KChIP2. Association with the full-size KChIPs is known to generate two prominent changes in the gating of Kv4 channels: slowing of inactivation and facilitation of recovery from inactivation. The newly identified two EF-hand-containing splicing variant produces both electrophysiological changes in its associated Kv4.3 channel in Xenopus oocytes. The authors also took advantage of this small peptide to show that mutations in the EF-hands selectively eliminate the KChIP2d-induced change in inactivation kinetics, but not recovery from inactivation.

This study has provided new insights into the structure and function of KChIP-Kv4 channel complexes. Structurally, the work demonstrated that the C-terminal half of the KChIP protein is sufficient for its interaction with Kv4 channel. Assuming that the two EF-hands are not directly involved in this interaction, the straightforward interpretation of the data is that the linker between the last two EF-hands and the C-terminal region after the last EF-hand would constitute the site for association with the channel protein. Functionally, Ca²⁺ binding to KChIP2d appears to relieve the KChIP-induced slowing of inactivation. A similar Ca²⁺-induced elimination of the slowing of inactivation is seen with Kv4-NCS-1 complexes (Nakamura et al. 2001). These results imply that intracellular Ca²⁺ concentration controls the gating of Kv4-KChIP or Kv4-NCS-1 complexes, resulting in the change in the time course of action potentials. However, it is important to note that the observed Ca²⁺-induced change in the channel gating may not be the only effect that Ca²⁺ produces on Kv4-KChIP complexes in a more physiological context.

The studies on KChIPs and their related proteins have yielded some surprising findings. These identified mechanisms may be important for fine tuning of excitability, as well as the link between excitability and other cellular functions. Over the next few years, we may not only see more surprises, but also be required to evaluate the physiological significance of the findings obtained.