MKRN1 induces degradation of West Nile virus capsid protein by functioning as an E3 ligase.
West Nile virus capsid protein (WNVCp) displays pathogenic toxicity via the apoptotic pathway. However, a cellular mechanism protective against this toxic effect has not been observed so far. Here, we identified Makorin ring finger protein 1 (MKRN1) as a novel E3 ubiquitin ligase for WNVCp. The cytotoxic effects of WNVCp as well as its expression levels were inhibited in U2OS cells that stably expressed MKRN1. Immunoprecipitation analyses revealed an interaction between MKRN1 and WNVCp. Domain analysis indicated that the C terminus of MKRN1 and the N terminus of WNVCp were required for the interaction. MKRN1 could induce WNVCp ubiquitination and degradation in a proteasome-dependent manner. Interestingly, the WNVCp mutant with amino acids 1 to 105 deleted WNVCp was degraded by MKRN1, whereas the mutant with amino acids 1 to 90 deleted was not. When three lysine sites at positions 101, 103, and 104 of WNVCp were replaced with alanine, MKRN1-mediated ubiquitination and degradation of the mutant were significantly inhibited, suggesting that these sites are required for the ubiquitination. Finally, U2OS cell lines stably expressing MKRN1 were resistant to cytotoxic effects of WNV. In contrast, cells depleted of MKRN1 were more susceptible to WNVCp cytotoxicity. Confirming this, overexpression of MKRN1 significantly reduced, but depletion of MKRN1 increased, WNV proliferation in 293T cells. Taken together, our results suggest that MKRN1 can protect cells from WNV by inducing WNVCp degradation.
Department of Biotechnology and Bioengineering, Sungkyunkwan University, Suwon 440-746, South Korea.
Aram
A
Ko
Eun-Woo
EW
Lee
Jung-Yong
JY
Yeh
Mi-Ran
MR
Yang
Wonkyung
W
Oh
Jin-San
JS
Moon
Jaewhan
J
Song
D019943:Amino Acid Substitution; D001665:Binding Sites; D036022:Capsid Proteins / Q000378:metabolism* / Q000502:physiology; D045744:Cell Line, Tumor; D006801:Humans; D008239:Lysine; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex; D012261:Ribonucleoproteins / Q000235:genetics / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*; D054875:Ubiquitination; D014902:West Nile virus / Q000737:chemistry / Q000472:pathogenicity*
D036022:Capsid Proteins; C410808:Makorin ring finger protein 1; D009419:Nerve Tissue Proteins; D012261:Ribonucleoproteins; D044767:Ubiquitin-Protein Ligases; D046988:Proteasome Endopeptidase Complex; D008239:Lysine