Antibody RING-Mediated Destruction of Endogenous Proteins

Summary To understand gene function, the encoding DNA or mRNA transcript can be manipulated and the consequences observed. However, these approaches do not have a direct effect on the protein product of the gene, which is either permanently abrogated or depleted at a rate defined by the half-life of the protein. We therefore developed a single-component system that could induce the rapid degradation of the specific endogenous protein itself. A construct combining the RING domain of ubiquitin E3 ligase RNF4 with a protein-specific camelid nanobody mediates target destruction by the ubiquitin proteasome system, a process we describe as antibody RING-mediated destruction (ARMeD). The technique is highly specific because we observed no off-target protein destruction. Furthermore, bacterially produced nanobody-RING fusion proteins electroporated into cells induce degradation of target within minutes. With increasing availability of protein-specific nanobodies, this method will allow rapid and specific degradation of a wide range of endogenous proteins.
Graphical Abstract
Highlights • Antibody RING-mediated destruction (ARMeD) targets endogenous proteins for degradation • ARMeD is mediated by a nanobody fused to the RING domain of ubiquitin E3 ligase RNF4 • Nanobody-RING fusions introduced into cells degrade target proteins within minutes
To study gene function, a single-component system that induces rapid degradation of the protein product of the gene was developed. Described as antibody RING-mediated destruction (ARMeD), a nanobody fused to the RING domain of ubiquitin E3 ligase RNF4 mediates degradation of the target protein by the ubiquitin proteasome system.