High spatial density is associated with non-conducting Kv channels from two families

Ion channels are well known for their ability to regulate the cell membrane potential. However, many ion channels also have functions that do not involve ion conductance. Kv2 channels are one family of ion channels whose non-conducting functions are central to mammalian cell physiology. Kv2.1 and Kv2.2 channels form stable contact sites between the endoplasmic reticulum and plasma membrane via an interaction with endoplasmic reticulum resident proteins. To perform this structural role, Kv2 channels are expressed at extremely high densities on the plasma membranes of many cell types, including central pyramidal neurons, α-motoneurons, and smooth muscle cells. Research from our lab and others has shown that the majority of these plasma membrane Kv2.1 channels do not conduct potassium in response to depolarization. The mechanism of this channel silencing is unknown but is thought to be dependent on channel density in the membrane. Furthermore, the prevalence of a non-conducting population of Kv2.2 channels has not been directly tested. In this work we make improved measurements of the numbers of conducting and non-conducting Kv2.1 channels expressed in HEK293 cells and expand the investigation of non-conducting channels to three additional Kv α-subunits: Kv2.2, Kv1.4, and Kv1.5. By comparing the numbers of gating and conducting channels in individual HEK293 cells, we found that on average, only 50% of both Kv2.1 and Kv2.2 channels conducted potassium and, as previously suggested, that fraction decreased with increased channel density in the plasma membrane. At the highest spatial densities tested, which are comparable with those found at Kv2 clusters in situ, only 20% of Kv2.1 and Kv2.2 channels conducted potassium. We also show for the first time that Kv1.4 and Kv1.5 exhibit density-dependent silencing, suggesting that this phenomenon has an underlying mechanism that is shared by Kv channels from multiple families.