Cullin 7 mediates proteasomal and lysosomal degradations of rat Eag1 potassium channels

Sci Rep. 2017 Jan 18:7:40825. doi: 10.1038/srep40825.

Abstract

Mammalian Eag1 (Kv10.1) potassium (K+) channels are widely expressed in the brain. Several mutations in the gene encoding human Eag1 K+ channel have been associated with congenital neurodevelopmental anomalies. Currently very little is known about the molecules mediating protein synthesis and degradation of Eag1 channels. Herein we aim to ascertain the protein degradation mechanism of rat Eag1 (rEag1). We identified cullin 7 (Cul7), a member of the cullin-based E3 ubiquitin ligase family, as a novel rEag1 binding partner. Immunoprecipitation analyses confirmed the interaction between Cul7 and rEag1 in heterologous cells and neuronal tissues. Cul7 and rEag1 also exhibited significant co-localization at synaptic regions in neurons. Over-expression of Cul7 led to reduced protein level, enhanced ubiquitination, accelerated protein turn-over, and decreased current density of rEag1 channels. We provided further biochemical and morphological evidence suggesting that Cul7 targeted endoplasmic reticulum (ER)- and plasma membrane-localized rEag1 to the proteasome and the lysosome, respectively, for protein degradation. Cul7 also contributed to protein degradation of a disease-associated rEag1 mutant. Together, these results indicate that Cul7 mediates both proteasomal and lysosomal degradations of rEag1. Our findings provide a novel insight to the mechanisms underlying ER and peripheral protein quality controls of Eag1 channels.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Membrane / metabolism
  • Cullin Proteins / genetics
  • Cullin Proteins / metabolism*
  • Cycloheximide / pharmacology
  • Endoplasmic Reticulum / metabolism
  • Ether-A-Go-Go Potassium Channels / genetics
  • Ether-A-Go-Go Potassium Channels / metabolism*
  • HEK293 Cells
  • Humans
  • Leupeptins / pharmacology
  • Lysosomes / metabolism*
  • Neurons / metabolism
  • Proteasome Endopeptidase Complex / chemistry
  • Proteasome Endopeptidase Complex / metabolism*
  • Protein Binding
  • Protein Stability / drug effects
  • Proteolysis / drug effects
  • Rats

Substances

  • Cullin Proteins
  • Ether-A-Go-Go Potassium Channels
  • Kcnh1 protein, rat
  • Leupeptins
  • Cycloheximide
  • Proteasome Endopeptidase Complex
  • benzyloxycarbonylleucyl-leucyl-leucine aldehyde