Discovery of cellular substrates of human RNA decapping enzyme Dcp2 using a stapled bicyclic peptide inhibitor

Dcp2 is an important enzyme that controls the stability of a subset of human cellular RNAs encoding functions such as transcription and immune responses. While >1800 Dcp2 substrates have been identified in human cells, compensatory gene expression changes secondary to genetic ablation of the DCP2 gene have complicated a complete mapping of its regulome. Cellpermeable, selective chemical ligands of Dcp2 could facilitate development of improved tools for elucidation of its function and cellular specificity. Here, we report the selection of a bicyclic peptide ligand of human Dcp2 from chemically cyclized phageencoded libraries, and evaluation of its affinity and selectivity for Dcp2 in vitro and in cells. Quantitative reverse transcription PCR (qRT-PCR) combined with a splinted ligation assay revealed that this stapled bicyclic peptide inhibits Dcp2 decapping activity toward specific RNA substrates inside human cells. We further demonstrate that CP21 increases formation of P-bodies, cellular liquid condensates enriched in intermediates of cellular RNA decay, in the same manner as deletion or mutation of Dcp2. Collectively, these results demonstrate development of a selective ligand of Dcp2 that modulates some of its functions inside cells. In the future, this molecule – and, more broadly, stapled peptide selection – may find utility in reverse chemical genetics to dissect the specificities of coexistent cellular RNA decay pathways.