ZIP-ZnT1 complexes mediate a local Zn2+-cycle regulating neuronal Zn²⁺ transport

Abstract ZnT1 is the main transporter mediating Zn²⁺ efflux from the cytoplasm to the extracellular space. Paradoxically, due to relatively low Zn 2+ affinity of ZnT1, physiological levels of cytoplasmic free Zn 2+ are seemingly insufficient to support ZnT1 function. However, physiological modulation of NMDA receptor responses by synaptically released Zn 2+ required ZnT1 activity, accompanied by a Zn 2+ rise in the postsynaptic neurons. We hypothesized that Zn 2+ import from the extracellular space could generate intracellular Zn 2+ microdomains in the vicinity of ZnT1 to enable its activity, thus forming a localized “Zn 2+ -cycle”. To identify the functional machinery that may drive this process, we investigated the expression and function of ZIP proteins that import Zn 2+ into the neuronal cytoplasm. We focused on the dorsal cochlear nucleus (DCN) and hippocampus, regions where synaptic Zn 2+ modulates postsynaptic NMDA responses and synaptic plasticity. We demonstrate that ZIP3 is expressed on DCN cartwheel cells and required for postsynaptic Zn 2+ influx. Importantly, ZIP3 physically interacts with ZnT1, suggesting that it is an integral component of the synaptic Zn 2+ -cycle machinery. In the hippocampus, ZIP1, but not ZIP3, mediates Zn 2+ import into postsynaptic CA3 cells and we find that ZIP1 interacts with ZnT1 in this brain region. Importantly, Zn 2+ efflux rates are enhanced in SH-SY5Y cells co-expressing ZnT1 and either ZIP3 or ZIP1, compared to rates in cells expressing ZnT1 alone. Our findings indicate that a Zn 2+ -cycle, mediated by ZIP-ZnT1 complexes, establishes local intracellular Zn 2+ microdomains that facilitate ZnT1-dependent Zn 2+ efflux, which, in turn, modulates synaptic signaling.