A programmable targeted protein-degradation platform for versatile applications in mammalian cells and mice.

Myriad physiological and pathogenic processes are governed by protein levels and modifications. Controlled protein activity perturbation is essential to studying protein function in cells and animals. Based on Trim-Away technology, we screened for truncation variants of E3 ubiquitinase Trim21 with elevated efficiency (ΔTrim21) and developed multiple ΔTrim21-based targeted protein-degradation systems (ΔTrim-TPD) that can be transfected into host cells. Three ΔTrim-TPD variants are developed to enable chemical and light-triggered programmable activation of TPD in cells and animals. Specifically, we used ΔTrim-TPD for (1) red-light-triggered inhibition of HSV-1 virus proliferation by degrading the packaging protein gD, (2) for chemical-triggered control of the activity of Cas9/dCas9 protein for gene editing, and (3) for blue-light-triggered degradation of two tumor-associated proteins for spatiotemporal inhibition of melanoma tumor growth in mice. Our study demonstrates that multiple ΔTrim21-based controllable TPD systems provide powerful tools for basic biology research and highlight their potential biomedical applications.

Silencing GnT-V reduces oxaliplatin chemosensitivity in human colorectal cancer cells through N-glycan alteration of organic cation transporter member 2.

Organic cation transporter member 2 (OCT2) is an N-glycosylated transporter that has been shown to be closely associated with the transport of antitumor drugs. Oxaliplatin, a platinum-based drug, is used for the chemotherapy of colorectal cancer (CRC). However, oxaliplatin resistance is a major challenge in the treatment of advanced CRC. The aim of the present study was to better understand the mechanism underlying the chemosensitivity of CRC cells to oxaliplatin. The present study describes a potential novel strategy for enhancing oxaliplatin sensitivity involving the glycosylation of this drug transporter, specifically the modification of ß-1,6-N-acetylglucosamine (GlcNAc) residues by N-acetylglucosaminyltransferase V (GnT-V). The results revealed that the downregulation of GnT-V inhibited the oxaliplatin chemosensitivity of CW-2 cells. Furthermore, the knockdown of GnT-V caused a marked reduction in the presence of ß-1,6-GlcNAc structures on OCT2 and decreased the localization of OCT2 in the cytomembrane, which were associated with a reduced uptake of oxaliplatin in wild-type and oxaliplatin-resistant CW-2 cells. Overall, the study provides novel insights into the molecular mechanism by which GnT-V regulates the chemosensitivity to oxaliplatin, which involves the modulation of the drug transporter OCT2 by N-glycosylation in CRC cells.