Cell surface targeting and clustering interactions between heterologously expressed PSD-95 and the Shal voltage-gated potassium channel, Kv4.2.
Kv4.2 is a voltage-gated potassium channel that is critical in controlling the excitability of myocytes and neurons. Processes that influence trafficking and surface distribution patterns of Kv4.2 will affect its ability to contribute to cellular functions. The scaffolding/clustering protein PSD-95 regulates trafficking and distribution of several receptors and Shaker family Kv channels. We therefore investigated whether the C-terminal valine-serine-alanine-leucine (VSAL) of Kv4.2 is a novel binding motif for PSD-95. By using co-immunoprecipitation assays, we determined that full-length Kv4.2 and PSD-95 interact when co-expressed in mammalian cell lines. Mutation analysis in this heterologous expression system showed that the VSAL motif of Kv4.2 is necessary for PSD-95 binding. PSD-95 increased the surface expression of Kv4.2 protein and caused it to cluster, as shown by deconvolution microscopy and biotinylation assays. Deleting the C-terminal VSAL motif of Kv4.2 eliminated these effects, as did substituting a palmitoylation-deficient PSD-95 mutant. In addition to these effects of PSD-95 on Kv4.2 distribution, the channel itself promoted redistribution of PSD-95 to the cell surface in the heterologous expression system. This work represents the first evidence that a member of the Shal subfamily of Kv channels can bind to PSD-95, with functional consequences.
Wong
Wei
W
Division of Cellular and Molecular Biology, Toronto Western Research Institute, University Health Network, Toronto, Ontario M5T 2S8, Canada.
Newell
Evan W
EW
Jugloff
Denis G M
DG
Jones
Owen T
OT
Schlichter
Lyanne C
LC
D020816:Amino Acid Motifs; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D017931:DNA Primers; D000076122:Disks Large Homolog 4 Protein; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D015640:Ion Channel Gating; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015221:Potassium Channels / Q000378:metabolism*; D024642:Potassium Channels, Voltage-Gated; D011485:Protein Binding; D051381:Rats; D051672:Shal Potassium Channels
D017931:DNA Primers; D000076122:Disks Large Homolog 4 Protein; C501449:Dlg4 protein, rat; D047908:Intracellular Signaling Peptides and Proteins; C497018:KCND2 protein, human; C497020:Kcnd2 protein, rat; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated; D051672:Shal Potassium Channels; C033652:postsynaptic density proteins