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.
Highlights
Normal cellular functions, and emerging evidence supports the view that imbalances between the quality control system and the degradation machinery of ion channels can result in many human diseases[18,25,26]
In order to search for rEag1-interacting proteins that may be involved in the regulation of protein ubiquitination and degradation, we carried out yeast two-hybrid screening of a rat brain cDNA library by using a bait comprising a cytoplasmic carboxyl-terminal region of rEag[1]
One of the positive clones isolated by the screening was Cul[7]
Summary
Normal cellular functions, and emerging evidence supports the view that imbalances between the quality control system and the degradation machinery of ion channels can result in many human diseases[18,25,26]. Very little is known about the molecular regulation of the synthesis and degradation of Eag K+ channels. To further address the potential physiological and pathological roles of Eag channels in the mammalian brain, we aim to ascertain the protein degradation mechanism of rat Eag[1] (rEag1) proteins. Morphological, and electrophysiological methods to characterize this novel protein interaction between Cul[7] and rEag[1], we conclude that Cul[7] plays an important role in the molecular machinery dictating the ubiquitin-dependent regulation of rEag[1] K+ channels in both the ER and the plasma membrane
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
