Small molecule WDR5 inhibitors down-regulate lncRNA expression.

WD repeat domain 5 (WDR5) plays an important role as a scaffold protein in both protein-protein and RNA-protein complexes involved in epigenetic gene regulation. In particular, some of these lncRNAs were reported to regulate the expression of genes in cis as well as themselves through binding WDR5. In this report, we investigate the two known binding sites of WDR5 in relation to lncRNA binding and expression. The WBM binding site mediates both protein-protein and lncRNA-protein interactions while the WIN site, which is on the opposite side of the protein, is only known to mediate protein-protein interactions. To dissect the function of different binding sites on WDR5, we characterized them with selective peptide ligands using fluorescence polarization and used these to demonstrate the selectivity of small molecule inhibitors of these two major binding sites. RNA immunoprecipitation experiments were performed to show that lncRNA-WDR5 complex formation could be interrupted using a WBM site inhibitor. Finally, we demonstrated that WDR5 regulated lncRNAs are down regulated with different sensitivity toward the corresponding inhibitors, demonstrating the potential of targeting lncRNA-protein interactions to reduce oncogenic lncRNA expression.

Macrocyclic peptides as inhibitors of WDR5-lncRNA interactions.

WDR5 is an adaptor protein involved in the regulation of various epigenetic modifier complexes. Various inhibitors have been described but only as inhibitors of its protein-protein interactions. Here we describe peptidic macrocycles that act as inhibitors of the interaction between WDR5 and long non-coding RNAs. The findings provide a new strategy to modulate the biological function of WDR5 as an RNA binding epigenetic regulator.

Rationally designed stapled peptides allosterically inhibit PTBP1-RNA-binding.

The diverse role of the splicing factor PTBP1 in human cells has been widely studied and was found to be a driver for several diseases. PTBP1 binds RNA through its RNA-recognition motifs which lack obvious pockets for inhibition. A unique transient helix has been described to be part of its first RNA-recognition motif and to be important for RNA binding. In this study, we further confirmed the role of this helix and envisioned its dynamic nature as a unique opportunity to develop stapled peptide inhibitors of PTBP1. The peptides were found to be able to inhibit RNA binding via fluorescence polarization assays and directly occupy the helix binding site as observed by protein crystallography. These cell-permeable inhibitors were validated in cellulo to alter the regulation of alternative splicing events regulated by PTBP1. Our study demonstrates transient secondary structures of a protein can be mimicked by stapled peptides to inhibit allosteric mechanisms.

Solid-Phase Peptide Modification via Deaminative Photochemical Csp3 -Csp3 Bond Formation Using Katritzky Salts.

Introduction of unnatural amino acids can significantly improve the binding affinity and stability of peptides. Commercial availability of such amino acids is limited, and their synthesis is a long and tedious process. We here describe a method that allows the functionalization of peptides directly on solid-support by converting lysine residues to Katritzky salts, and subjecting them to a photochemical Giese reaction under mild reaction conditions. The method avoids the need for amino acid synthesis and instead offers a late-stage modification route for rapid peptide diversification. While numerous modification approaches at the lysine amine have been described, this work provides the first example of deaminative functionalization of peptides at lysine. The two-step protocol is compatible with various substrates, lysine analogues, resins, and all proteinogenic amino acids. Finally, by leveraging solid-phase modification, this protocol facilitates the functionalization of longer peptides as was demonstrated using biologically relevant peptides of up to 15 amino acids.