Migrating transformed MDCK cells are able to structurally polarize a voltage-activated K+ channel.
Cell migration of transformed renal epithelial cells (MDCK-F) depends-in addition to cytoskeletal mechanisms-on the polarized activity of a Ca2+-sensitive K+ channel in the rear part of the cells. However, because of the lack of specific markers for this channel we are not able to determine whether a polarized distribution of the channel protein underlies its functional polarization. To determine whether the migrating MDCK-F cells have retained the ability to target K+ channels to distinct membrane areas we stably transfected the cells with the voltage-dependent K+ channel Kv1.4. Stable expression and insertion into the plasma membrane could be shown by reverse transcription-PCR, genomic PCR, Western blot, and patch-clamp techniques, respectively. The distribution of Kv1.4 was assessed with indirect immunofluorescence by using conventional and confocal microscopy. These experiments revealed that Kv1.4 is expressed only in transfected cells where it elicits the typical voltage-dependent, rapidly inactivating K+ current. The Kv1.4 protein is clustered at the leading edge of protruding lamellipodia of migrating MDCK-F cells. This characteristic distribution of Kv1.4 provides strong evidence that migrating MDCK-F cells are able to insert ion channels into the plasma membrane in an asymmetric way, which reflects the polarization of migrating cells in the plane of movement. These findings suggest that not only epithelial cells and nerve cells, but also migrating cells, can create functionally distinct plasma membrane areas.
Reinhardt
J
J
Physiologisches Institut, Universität Würzburg, Röntgenring 9, D-97070 Würzburg, Germany. Juergen.Reinhardt@quickmail.yale.edu
Golenhofen
N
N
Pongs
O
O
Oberleithner
H
H
Schwab
A
A
D000818:Animals; D002460:Cell Line; D002465:Cell Movement; D016764:Cell Polarity; D004285:Dogs; D019084:Fluorescent Antibody Technique, Indirect; D051665:Kv1.4 Potassium Channel; D008564:Membrane Potentials; D018613:Microscopy, Confocal; D011188:Potassium / Q000502:physiology*; D015221:Potassium Channels / Q000378:metabolism*; D024642:Potassium Channels, Voltage-Gated; D014162:Transfection
D051665:Kv1.4 Potassium Channel; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated; D011188:Potassium