Identification of MKRN1 as a second E3 ligase for Eag1 potassium channels reveals regulation via differential degradation

Abstract

Mutations in the human gene encoding the neuron-specific Eag1 voltage-gated K + channel are associated with neurodevelopmental diseases, indicating an important role of Eag1 during brain development. A disease-causing Eag1 mutation is linked to decreased protein stability that involves enhanced protein degradation by the E3 ubiquitin ligase cullin 7 (CUL7). The general mechanisms governing protein homeostasis of plasma membrane- and endoplasmic reticulum (ER)-localized Eag1 K + channels, however, remain unclear. By using yeast two-hybrid screening, we identified another E3 ubiquitin ligase, makorin ring finger protein 1 (MKRN1), as a novel binding partner primarily interacting with the carboxyl-terminal region of Eag1. MKRN1 mainly interacts with ER-localized immature core-glycosylated, as well as nascent nonglycosylated, Eag1 proteins. MKRN1 promotes polyubiquitination and ER-associated proteasomal degradation of immature Eag1 proteins. Although both CUL7 and MKRN1 contribute to ER quality control of immature core-glycosylated Eag1 proteins, MKRN1, but not CUL7, associates with and promotes degradation of nascent, nonglycosylated Eag1 proteins at the ER. In direct contrast to the role of CUL7 in regulating both ER and peripheral quality controls of Eag1, MKRN1 is exclusively responsible for the early stage of Eag1 maturation at the ER. We further demonstrated that both CUL7 and MKRN1 contribute to protein quality control of additional disease-causing Eag1 mutants associated with defective protein homeostasis. Our data suggest that the presence of this dual ubiquitination system differentially maintains Eag1 protein homeostasis and may ensure efficient removal of disease-associated misfolded Eag1 mutant channels.

Rare SLC13A1 variants associate with intervertebral disc disorder highlighting role of sulfate in disc pathology

“…For IDD these include SERPINA1 ( Serpin family A member 1 ), that encodes a serine protease inhibitor belonging to the serpin superfamily whose targets include elastase, plasmin, thrombin, trypsin, chymotrypsin, and plasminogen activator; the transcription factor MYPOP ( Myb-related transcription factor, partner of profilin ); CNNM2 ( Cyclin M2 ) that encodes a transmembrane protein involved in magnesium transport; FGFR3 ( Fibroblast growth factor receptor 3 ) and TGFA ( Transforming growth factor alpha ), both encoding growth factors involved in bone development; GFPT1 , encoding Glutamine fructose-6-phosphate amidotransferase 1 , which is the first and rate-limiting enzyme of the hexosamine biosynthetic pathway and has been linked to recessive congenital myasthenic syndrome and synthesis of proteoglycans 35 . Of the six novel dorsalgia genes, five associate more strongly with dorsalgia than IDD, including MKRN1 ( Makorin ring finger protein-1 ), an E3 ubiquitin ligase involved in protein homeostasis of Eag1 potassium channels 36 , EIF4E3 ( Eukaryotic translation initiation factor 4E family member 3 ) and SNRPC ( Small nuclear ribonucleprotein polypetide C ); both widely expressed in tissues and involved in mRNA translation. SIGLECL1 (Sialic acid-binding immunoglobulin-like lectin 12) encodes a cell surface protein of the Ig superfamily and is mainly expressed in the immune system 37 .…”

The MKRN1-BRAF exon4-exon9 fusion is a targetable oncogenic driver

“…MKRN1, also known as Makorin, is an E3 ubiquitin ligase. MKRN1 substrates include the EAG1 potassium channel (Fang et al, 2021), the catalytic subunit of telomerase, hTERT (Kim et al, 2005), as well as p53 and p21 (Lee et al, 2009). Experimental depletion of MKRN1 resulted in increased telomerase activity (Kim et al, 2005) and suppression of DNA damage induced cell death (Lee et al, 2009), either of which could confer additional advantages beyond activation of MEK/ERK signaling during cellular transformation or subsequent therapy.…”

The Kv10.1 Channel: A Promising Target in Cancer

“…Cul7 degrades both endoplasmic reticulum-localized immature and plasma membrane-localized matures Kv10.1 protein through the proteasomal and the lysosomal pathway, respectively [ 79 ]. In cooperation with this mechanism, the E3 ubiquitin ligase makorin ring finger protein 1 (MKRN1) has been described as another endoplasmic reticulum-localized mechanism of Kv10.1 degradation [ 80 ]. Both works demonstrated that Cul7 and MKRN1 are quality checkpoints of immature Kv10.1 channels.…”