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bioRxiv · 10.1101/2021.10.23.465516

Activity-dependent endoplasmic reticulum Ca2+ uptake depends on Kv2.1-mediated endoplasmic reticulum/plasma membrane junctions to promote synaptic transmission

Panzera, L. C.·Johnson, B.·Cho, I. H.·Tamkun, M. M.·Hoppa, M. B.
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Abstract

The endoplasmic reticulum (ER) forms a continuous and dynamic network throughout a neuron, extending from dendrites to axon terminals, and axonal ER dysfunction is implicated in several neurological disorders. In addition, tight junctions between the ER and plasma membrane (PM) are formed by several molecules including Kv2 channels, but the cellular functions of many ER-PM junctions remain unknown. Dynamic Ca2+ uptake into the ER during electrical activity plays an essential role in synaptic transmission as failure to allow rapid ER Ca2+ filling during stimulation activates stromal interaction molecule 1 (STIM1) and decreases both presynaptic Ca2+ influx and synaptic vesicle exocytosis. Our experiments demonstrate that Kv2.1 channels are necessary for enabling ER Ca2+ uptake during electrical activity as genetic depletion of Kv2.1 rendered both the somatic and axonal ER unable to accumulate Ca2+ during electrical stimulation. Moreover, our experiments show that the loss of Kv2.1 in the axon impairs synaptic vesicle fusion during stimulation via a mechanism unrelated to modulation of membrane voltage. Thus, our data demonstrate that the non-conducting role of Kv2.1 in forming stable junctions between the ER and PM via ER VAMP-associated protein (VAP) binding couples ER Ca2+ uptake with electrical activity. Our results further suggest that Kv2.1 has a critical function in neuronal cell biology for Ca2+-handling independent of voltage and reveals a novel and critical pathway for maintaining ER lumen Ca2+ levels and efficient neurotransmitter release. Taken together these findings reveal an essential non-classical role for both Kv2.1 and the ER-PM junctions in synaptic transmission. SignificanceThe endoplasmic reticulum (ER) extends throughout the neuron as a continuous organelle, and its dysfunction is associated with several neurological disorders. During electrical activity, the ER takes up Ca2+ from the cytosol which has been shown to support synaptic transmission. This close choreography of ER Ca2+ uptake with electrical activity suggests functional coupling of the ER to sources of voltage-gated Ca2+ entry through an unknown mechanism. Here we report a non-conducting role for Kv2.1 through its ER binding domain that is necessary for ER Ca2+ uptake during neuronal activity. Loss of Kv2.1 profoundly disables neurotransmitter release without altering presynaptic voltage suggesting that Kv2.1-mediated signaling hubs play an important neurobiological role in Ca2+ handling and synaptic transmission independent of ion conduction.

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Panzera, L. C., Johnson, B., Cho, I. H., Tamkun, M. M., Hoppa, M. B.. 2021-10-23. Activity-dependent endoplasmic reticulum Ca2+ uptake depends on Kv2.1-mediated endoplasmic reticulum/plasma membrane junctions to promote synaptic transmission. https://doi.org/10.1101/2021.10.23.465516

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