Loss of peptide: <i>N</i> -glycanase causes proteasome dysfunction mediated by a sugar-recognizing ubiquitin ligase

Abstract

Mutations in the human peptide:<i>N</i>-glycanase gene (<i>NGLY1</i>), which encodes a cytosolic de-<i>N</i>-glycosylating enzyme, cause a congenital autosomal recessive disorder. In rodents, the loss of <i>Ngly1</i> results in severe developmental delay or lethality, but the underlying mechanism remains unknown. In this study, we found that deletion of <i>Fbxo6</i> (also known as <i>Fbs2</i>), which encodes a ubiquitin ligase subunit that recognizes glycoproteins, rescued the lethality-related defects in <i>Ngly1</i>-KO mice. In <i>NGLY1</i>-KO cells, FBS2 overexpression resulted in the substantial inhibition of proteasome activity, causing cytotoxicity. Nuclear factor, erythroid 2-like 1 (NFE2L1, also known as NRF1), an endoplasmic reticulum-associated transcriptional factor involved in expression of proteasome subunits, was also abnormally ubiquitinated by SCF^FBS2 in <i>NGLY1</i>-KO cells, resulting in its retention in the cytosol. However, the cytotoxicity caused by FBS2 was restored by the overexpression of "glycan-less" NRF1 mutants, regardless of their transcriptional activity, or by the deletion of <i>NRF1</i> in <i>NGLY1</i>-KO cells. We conclude that the proteasome dysfunction caused by the accumulation of <i>N</i>-glycoproteins, primarily NRF1, ubiquitinated by SCF^FBS2 accounts for the pathogenesis resulting from NGLY1 deficiency.