The C-terminus SH3-binding domain of Kv1.3 is required for the actin-mediated immobilization of the channel via cortactin.
Kv1.3 channels play a pivotal role in the activation and migration of T-lymphocytes. These functions are accompanied by the channels' polarization, which is essential for associated downstream events. However, the mechanisms that govern the membrane movement of Kv1.3 channels remain unclear. F-actin polymerization occurs concomitantly to channel polarization, implicating the actin cytoskeleton in this process. Here we show that cortactin, a factor initiating the actin network, controls the membrane mobilization of Kv1.3 channels. FRAP with EGFP-tagged Kv1.3 channels demonstrates that knocking down cortactin decreases the actin-based immobilization of the channels. Using various deletion and mutation constructs, we show that the SH3 motif of Kv1.3 mediates the channel immobilization. Proximity ligation assays indicate that deletion or mutation of the SH3 motif also disrupts interaction of the channel with cortactin. In T-lymphocytes, the interaction between HS1 (the cortactin homologue) and Kv1.3 occurs at the immune synapse and requires the channel's C-terminal domain. These results show that actin dynamics regulates the membrane motility of Kv1.3 channels. They also provide evidence that the SH3 motif of the channel and cortactin plays key roles in this process.
Division of Nephrology and Hypertension, Department of Internal Medicine, University of Cincinnati, Cincinnati, OH 45267 Department of Biophysics and Cell Biology, University of Debrecen, 4032 Debrecen, Hungary.
Peter
P
Hajdu
Division of Nephrology and Hypertension, Department of Internal Medicine, University of Cincinnati, Cincinnati, OH 45267.
Geoffrey V
GV
Martin
Division of Nephrology and Hypertension, Department of Internal Medicine, University of Cincinnati, Cincinnati, OH 45267.
Ameet A
AA
Chimote
Division of Nephrology and Hypertension, Department of Internal Medicine, University of Cincinnati, Cincinnati, OH 45267 Department of Biophysics and Cell Biology, University of Debrecen, 4032 Debrecen, Hungary.
Orsolya
O
Szilagyi
Department of Bioengineering and Bioinformatics, Nagaoka University of Technology, Nagaoka 940-2137, Japan.
Koichi
K
Takimoto
Division of Nephrology and Hypertension, Department of Internal Medicine, University of Cincinnati, Cincinnati, OH 45267 laura.conforti@uc.edu.
Laura
L
Conforti
D008841:Actin Cytoskeleton / Q000378:metabolism*; D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D001665:Binding Sites; D001798:Blood Proteins / Q000378:metabolism; D051356:Cortactin / Q000378:metabolism*; D036681:Fluorescence Recovery After Photobleaching; D057809:HEK293 Cells; D006801:Humans; D054992:Immunological Synapses / Q000378:metabolism; D007700:Kinetics; D051664:Kv1.3 Potassium Channel / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D021381:Protein Transport; D013601:T-Lymphocytes / Q000378:metabolism; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D001798:Blood Proteins; D051356:Cortactin; C061909:HCLS1 protein, human; D051664:Kv1.3 Potassium Channel