Identification of the HECT E3 ligase UBR5 as a regulator of MYC degradation using a CRISPR/Cas9 screen.

Scientific Reports
Lina SchukurClaudio R Thoma

Abstract

MYC oncoprotein is a multifunctional transcription factor that regulates the expression of a large number of genes involved in cellular growth, proliferation and metabolism. Altered MYC protein level lead to cellular transformation and tumorigenesis. MYC is deregulated in > 50% of human cancers, rendering it an attractive drug target. However, direct inhibition of this class of proteins using conventional small molecules is challenging due to their intrinsically disordered state. To discover novel posttranslational regulators of MYC protein stability and turnover, we established a genetic screen in mammalian cells by combining a fluorescent protein-based MYC abundance sensor, CRISPR/Cas9-based gene knockouts and next-generation sequencing. Our screen identifies UBR5, an E3 ligase of the HECT-type family, as a novel regulator of MYC degradation. Even in the presence of the well-described and functional MYC ligase, FBXW7, UBR5 depletion leads to accumulation of MYC in cells. We demonstrate interaction of UBR5 with MYC and reduced K48-linked ubiquitination of MYC upon loss of UBR5 in cells. Interestingly, in cancer cell lines with amplified MYC expression, depletion of UBR5 resulted in reduced cell survival, as a consequence of MY...Continue Reading

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Citations

Mar 30, 2021·EMBO Reports·Kevin G Mark, Michael Rape

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Methods Mentioned

BETA
ubiquitination
genetic modification
deubiquitinases
gene knockout
Confocal microscopy
FACS
flow cytometry
chip
transfection
neddylation

Software Mentioned

flowjo
DRIVE
ImageJ
RSA
CV7000
FACS
Bioconductor package edgeR
edgeR

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