Selective Interaction of Syntaxin 1A with KCNQ2: Possible Implications for Specific Modulation of Presynaptic Activity

KCNQ2/KCNQ3 channels are the molecular correlates of the neuronal M-channels, which play a major role in the control of neuronal excitability. Notably, they differ from homomeric KCNQ2 channels in their distribution pattern within neurons, with unique expression of KCNQ2 in axons and nerve terminals. Here, combined reciprocal coimmunoprecipitation and two-electrode voltage clamp analyses in Xenopus oocytes revealed a strong association of syntaxin 1A, a major component of the exocytotic SNARE complex, with KCNQ2 homomeric channels resulting in a ∼2-fold reduction in macroscopic conductance and ∼2-fold slower activation kinetics. Remarkably, the interaction of KCNQ2/Q3 heteromeric channels with syntaxin 1A was significantly weaker and KCNQ3 homomeric channels were practically resistant to syntaxin 1A. Analysis of different KCNQ2 and KCNQ3 chimeras and deletion mutants combined with in-vitro binding analysis pinpointed a crucial C-terminal syntaxin 1A-association domain in KCNQ2. Pull-down and coimmunoprecipitation analyses in hippocampal and cortical synaptosomes demonstrated a physical interaction of brain KCNQ2 with syntaxin 1A, and confocal immunofluorescence microscopy showed high colocalization of KCNQ2 and syntaxin 1A at presynaptic varicosities. The selective interaction of syntaxin 1A with KCNQ2, combined with a numerical simulation of syntaxin 1A's impact in a firing-neuron model, suggest that syntaxin 1A's interaction is targeted at regulating KCNQ2 channels to fine-tune presynaptic transmitter release, without interfering with the function of KCNQ2/3 channels in neuronal firing frequency adaptation

Fast inactivation of a brain K+ channel composed of Kv1.1 and Kvbeta1.1 subunits modulated by G protein beta gamma subunits.

Modulation of A-type voltage-gated K+ channels can produce plastic changes in neuronal signaling. It was shown that the delayed-rectifier Kv1.1 channel can be converted to A-type upon association with Kvbeta1.1 subunits; the conversion is only partial and is modulated by phosphorylation and microfilaments. Here we show that, in Xenopus oocytes, expression of Gbeta1gamma2 subunits concomitantly with the channel (composed of Kv1.1 and Kvbeta1.1 subunits), but not after the channel's expression in the plasma membrane, increases the extent of conversion to A-type. Conversely, scavenging endogenous Gbetagamma by co-expression of the C-terminal fragment of the beta-adrenergic receptor kinase reduces the extent of conversion to A-type. The effect of Gbetagamma co-expression is occluded by treatment with dihydrocytochalasin B, a microfilament-disrupting agent shown previously by us to enhance the extent of conversion to A-type, and by overexpression of Kvbeta1.1. Gbeta1gamma2 subunits interact directly with GST fusion fragments of Kv1.1 and Kvbeta1.1. Co-expression of Gbeta1gamma2 causes co-immunoprecipitation with Kv1.1 of more Kvbeta1.1 subunits. Thus, we suggest that Gbeta1gamma2 directly affects the interaction between Kv1.1 and Kvbeta1.1 during channel assembly which, in turn, disrupts the ability of the channel to interact with microfilaments, resulting in an increased extent of A-type conversion