Recursive Editing improves homology-directed repair through retargeting of undesired outcomes.

CRISPR-Cas induced homology-directed repair (HDR) enables the installation of a broad range of precise genomic modifications from an exogenous donor template. However, applications of HDR in human cells are often hampered by poor efficiency, stemming from a preference for error-prone end joining pathways that yield short insertions and deletions. Here, we describe Recursive Editing, an HDR improvement strategy that selectively retargets undesired indel outcomes to create additional opportunities to produce the desired HDR allele. We introduce a software tool, named REtarget, that enables the rational design of Recursive Editing experiments. Using REtarget-designed guide RNAs in single editing reactions, Recursive Editing can simultaneously boost HDR efficiencies and reduce undesired indels. We also harness REtarget to generate databases for particularly effective Recursive Editing sites across the genome, to endogenously tag proteins, and to target pathogenic mutations. Recursive Editing constitutes an easy-to-use approach without potentially deleterious cell manipulations and little added experimental burden.

Translating from Proteins to Ribonucleic Acids for Ligand-binding Site Detection.

Identifying druggable ligand-binding sites on the surface of the macromolecular targets is an important process in structure-based drug discovery. Deep-learning models have been shown to successfully predict ligand-binding sites of proteins. As a step toward predicting binding sites in RNA and RNA-protein complexes, we employ three-dimensional convolutional neural networks. We introduce a dataset splitting approach to minimize structure-related bias in training data, and investigate the influence of protein-based neural network pre-training before fine-tuning on RNA structures. Models that were pre-trained on proteins considerably outperformed the models that were trained exclusively on RNA structures. Overall, 71 % of the known RNA binding sites were correctly located within 4 Šof their true centres.