UCSF

Long-term changes in synaptic strength are thought to underlie many forms of learning and memory. To achieve these changes in synaptic strength, neurons must be able to regulate the trafficking of neurotransmitter receptors to and from the synapse locally within dendrites. The dendritic trafficking of AMPAR-type glutamate receptors critically modulates synaptic strength, though the regulatory mechanisms are not fully understood. Here I describe a novel protein kinase D1 (PKD1)-dependent pathway that mediates the postsynaptic trafficking of AMPARs. Without functional PKD1, GluA2-containing AMPARs accumulated in endosomes, decreasing the number of extrasynaptic AMPARs. This is due to decreased GluA2 exocytosis, as knockdown of PKD1 reduced GluA2 recycling, but not uptake, in response to glutamate. PKD1 is in turn dually regulated by glutamate. We found that mGluRs control PKD1 activity, whereas NMDARs control PKD1 localization. Ca²⁺ influx through NMDARs caused movement of PKD1 from the dendritic cytoplasm to AMPAR-containing endosomes. PKD1 translocation is a multistep process requiring both its membrane-binding and kinase domains. DAG generated downstream of NMDARs recruited PKD1 to membranes, and PKD1 subsequently translocated to endosomes via its kinase domain. Mutating PKD1's activation loop serines to alanines (SSAA) disrupted endosomal localization, despite the fact that NMDARs do not induce PKD1 phosphorylation. I generated an ATP analog inhibitor-sensitive allele of PKD1 to determine if phosphoryl transfer activity is required for endosomal localization, but found inhibitor binding did not block PKD1 translocation. On the contrary, I found inhibitor binding rescued endosomal localization of the SSAA mutant, suggesting inhibitor binding induces the closed or "active" kinase conformation, and this conformation may be required for endosomal targeting. My results underscore the complexity with which PKD1 is regulated in neurons, and opens up the possibility that PKD1 localization to endosomes with and without concomitant kinase activation have different effects on AMPAR trafficking. Thus, PKD1 may be a critical node in a plasticity pathway that receives distinct synaptic signals from NMDARs and mGluRs and mediates changes in the availability of extrasynaptic AMPARs.