Inhibition of UBE2N promotes the clearance of mutant HTT (huntingtin) in HD knock-in mice

Accumulation of misfolded proteins leads to many neurodegenerative diseases that can be treated by lowering or removing mutant proteins. Huntington disease (HD) is characterized by the accumulation of ubiquitinated mutant HTT (huntingtin) in the central nervous system. Ubiquitination of the misfolded proteins, a common feature of the neurodegenerative diseases, is mediated by the different lysine residues on ubiquitin. We previously discovered that the age-dependent increase of UBE2N (ubiquitin conjugating enzyme E2 N) exacerbated the accumulation of misfolded HTT and amyloid proteins, accompanied by the elevation of K63 ubiquitination. Pharmacological inhibition of UBE2N could ameliorate the amyloid deposition. However, the effect of UBE2N suppression on HTT aggregate clearance has remained unknown. In the current work, we demonstrate that selectively suppressing UBE2N, with antisense oligonucleotides or small-molecular inhibitors, increased removal of HTT aggregates by proteasome degradation in the striatum of HD knock-in mice. We also identified two novel ubiquitin specific peptidases, USP29 and USP49, that participated in the clearance of HTT aggregates, via accelerating K48-mediated ubiquitin-proteasome function. Our findings provide a potential pharmacological approach to treat neurodegeneration caused by mutant HTT.Abbreviation: AD: Alzheimer disease; ALP: autophagy-lysosomal pathway; AMC: 7-Amino-4-Methylcoumarin; ASO: antisense oligonucleotide; Aβ: amyloid β; BafA1: bafilomycin A₁; DEG: differentially expressed gene; DMSO: dimethyl sulfoxide; E2: ubiquitin-conjugating; E3: ubiquitin-ligating; EGFP: enhanced green fluorescent protein; HD: Huntington disease; HTT: huntingtin; KI: knock-in; SDS: sodium dodecyl sulfate; UPS: ubiquitin-proteasome system; UBE2N: ubiquitin conjugating enzyme E2 N; USP: ubiquitin-specific peptidase.