The End of Protein Mislocalization

Degrons are minimal elements within substrate proteins that enable recognition by the degradative machinery. N-degrons and C-degrons that lie at protein termini have been shown to influence a wide variety of cellular functions. Here we focus on terminal degrons that play a regulatory role in protein quality control (PQC) mechanisms by mediating destruction of mislocalized proteins.

To characterize terminal degron motifs in human proteins in a systematic manner we utilized the GPS-peptidome approach. This technology is a hybrid of the Global Protein Stability (GPS) reporter system combined with a synthetic human peptidome library. GPS is based on a lentiviral construct encoding two fluorescent proteins: DsRed, which serves as an internal reference, and a GFP fusion peptide that is translated from an internal ribosome entry site (IRES). As both DsRed and the GFP fusion peptide are expressed from the same transcript, the GFP/DsRed ratio can be used as a readout of the effect of the fusion peptide on the stability of GFP.

Interestingly, using the GPS-peptidome approach we discover terminal degrons that function to preserve correct subcellular localization of proteins. We found that upon mislocalization, the terminal degron is exposed by proteases to enable recognition by E3 ligases and subsequent degradation of the mislocalized proteins. Altogether, our systematic approach helps to understand the mechanism of substrate selection by E3 ligases and its importance for proteostasis control.