The Axon–Dendrite Targeting of Kv3 (Shaw) Channels Is Determined by a Targeting Motif That Associates with the T1 Domain and Ankyrin G

Abstract

Kv3 (Shaw) channels regulate rapid spiking, transmitter release and dendritic integration of many central neurons. Crucial to functional diversity are the complex targeting patterns of channel proteins. However, the targeting mechanisms are not known. Here we report that the axon-dendrite targeting of Kv3.1 is controlled by a conditional interaction of a C-terminal axonal targeting motif (ATM) with the N-terminal T1 domain and adaptor protein ankyrin G. In cultured hippocampal neurons, although the two splice variants of Kv3.1, Kv3.1a and Kv3.1b, are differentially targeted to the somatodendritic and axonal membrane, respectively, the lysine-rich ATM is surprisingly common for both splice variants. The ATM not only directly binds to the T1 domain in a Zn 2ϩ -dependent manner, but also associates with the ankyrin-repeat domain of ankyrin G. However, the full-length channel proteins of Kv3.1b display stronger association to ankyrin G than those of Kv3.1a, suggesting that the unique splice domain at Kv3.1b C terminus influences ATM binding to T1 and ankyrin G. Because ankyrin G mainly resides at the axon initial segment, we propose that it may function as a barrier for axon-dendrite targeting of Kv3.1 channels. In support of this idea, disrupting ankyrin G function either by over-expressing a dominant-negative mutant or by siRNA knockdown decreases polarized axon-dendrite targeting of both Kv3.1a and Kv3.1b. We conclude that the conditional ATM masked by the T1 domain in Kv3.1a is exposed by the splice domain in Kv3.1b, and is subsequently recognized by ankyrin G to target Kv3.1b into the axon.

Clustering and Activity Tuning of Kv1 Channels in Myelinated Hippocampal Axons

“…cDNA Constructs and Transfection-The constructs Kv1.2HA, Kv1.2HAY458A, Kv1.2HAV499A, Kv␤2, YFP-Kv␤2, YFP-Kv1.2HA, and Kv3.1bHA were described previously (3,4). YFP-Kv1.2HAY458A was generated with a QuikChange strategy.…”

“…SigmaPlot10 was used for fitting. To obtain on , activation curves were fitted with a single exponential function raised to a power of 4, I(t) ϭ A(1 Ϫ exp(Ϫt/ on )) 4 . Clampfit10.0 was used for fitting.…”

“…Molecular mechanisms underlying polarized axon-dendrite targeting of Kv channels have been extensively studied (1)(2)(3)(4)(5)(6)(7)(8)(9)(10). In contrast, channel targeting in distinct membrane domains within axons is much less understood.…”

Activation of conventional kinesin motors in clusters by shaw voltage-gated potassium channels

“…Thus, clustered KIF5 motors can alter the behavior of microtubule tracks and indirectly affect the transport of other proteins. Therefore, our current and previous works showing Kv3 channels are linked to actin and microtubule cytoskeletons via ankyrin-G and KIF5, respectively (Xu et al, 2010;Xu et al, 2007), may shed light on this intriguing question. In particular, this work has raised a possibility that Kv3 channels may regulate axonal transport of other cargos via clustering kinesin-1, which is an interesting topic for further investigation.…”

“…Other cytoskeletal and associated proteins also regulate the specificity of kinesinmediated transport. For instance, ankyrin-G and actin filaments at the axon initial segment appear to be critical for maintaining the axon-dendrite polarity by regulating selective transport (Maniar et al, 2012;Xu et al, 2010;Xu et al, 2007;Sobotzik et al, 2009;Song et al, 2009). However, whether cargos themselves can play an important role during selective transport, knowing exactly which motor protein to ride, how to ride, and where to go, still remains unclear.…”