Post-synaptic density perturbs insulin-induced Kv1.3 channel modulation via a clustering mechanism involving the SH3 domain.
The olfactory bulb (OB) contains the highest concentration of the insulin receptor (IR) kinase in the central nervous system; however, its functional role and modulation in this region remains poorly understood. IR kinase contains a number of proline-rich motifs, making it an excellent candidate for modulation by SH(3) domain-containing adaptor proteins. Kv1.3, a voltage-gated Shaker potassium channel and tyrosine phosphorylation substrate of IR kinase, contains several proline-rich sequences and a canonical post-synaptic density 95 (PSD-95)/discs large/zO-1 domain (PDZ) recognition motif common to most Shaker family members. We sought to determine if a functional relationship existed between Kv1.3, IR kinase, and the SH(3)/PDZ adaptor protein PSD-95. Through patch-clamp electrophysiology, immunochemistry, and co-immunoprecipitation, we found that while Kv1.3 and PSD-95 alone interact via the canonical C-terminal PDZ recognition motif of the channel, this molecular site of interaction acts to cluster the channels but the PSD-95 SH(3)-guanylate kinase domain functionally modulates Kv1.3 activity via two proline-rich domains in its N- and C-terminal. Therefore, these data suggest that adaptor domains responsible for ion-channel clustering and functional modulation are not necessarily coupled. Moreover, IR kinase and Kv1.3 can only be co-immunoprecipitated in the presence of PSD-95 as the adapting linker. Functionally, insulin-dependent Kv1.3 phosphorylation that causes channel current suppression is blocked via interaction with the PSD-95 SH(3)-guanylate kinase domain. Because all the three proteins co-localize in multiple lamina of the OB that are known to be rich in synaptic connections, membrane excitability and synaptic transmission at critical locations in the OB have the capacity to be finely regulated.
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Department of Biological Science, Program in Neuroscience, Biomedical Research Facility, The Florida State University, Tallahassee, Florida, USA.
Fadool
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D020816:Amino Acid Motifs / Q000187:drug effects / Q000502:physiology; D002460:Cell Line; D000076122:Disks Large Homolog 4 Protein; D006801:Humans; D007328:Insulin / Q000378:metabolism / Q000502:physiology*; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism / Q000502:physiology*; D051664:Kv1.3 Potassium Channel / Q000378:metabolism* / Q000502:physiology; D008565:Membrane Proteins / Q000378:metabolism / Q000502:physiology*; D009419:Nerve Tissue Proteins / Q000378:metabolism / Q000502:physiology*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism / Q000502:physiology*; D011972:Receptor, Insulin / Q000378:metabolism / Q000502:physiology*; D018909:src Homology Domains / Q000502:physiology*
C501444:DLG4 protein, human; D000076122:Disks Large Homolog 4 Protein; D007328:Insulin; D047908:Intracellular Signaling Peptides and Proteins; D051664:Kv1.3 Potassium Channel; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; C033652:postsynaptic density proteins; D011972:Receptor, Insulin; D017346:Protein Serine-Threonine Kinases; C081827:insulin receptor serine kinase