Drag-and-drop genome insertion of large sequences without double-strand DNA cleavage using CRISPR-directed integrases.

Programmable genome integration of large, diverse DNA cargo without DNA repair of exposed DNA double-strand breaks remains an unsolved challenge in genome editing. We present programmable addition via site-specific targeting elements (PASTE), which uses a CRISPR-Cas9 nickase fused to both a reverse transcriptase and serine integrase for targeted genomic recruitment and integration of desired payloads. We demonstrate integration of sequences as large as ~36 kilobases at multiple genomic loci across three human cell lines, primary T cells and non-dividing primary human hepatocytes.

Programmable eukaryotic protein synthesis with RNA sensors by harnessing ADAR.

Programmable approaches to sense and respond to the presence of specific RNAs in biological systems have broad applications in research, diagnostics, and therapeutics. Here we engineer a programmable RNA-sensing technology, reprogrammable ADAR sensors (RADARS), which harnesses RNA editing by adenosine deaminases acting on RNA (ADAR) to gate translation of a cargo protein by the presence of endogenous RNA transcripts. Introduction of a stop codon in a guide upstream of the cargo makes translation contingent on binding of an endogenous transcript to the guide, leading to ADAR editing of the stop codon and allowing translational readthrough.

Dynamic postnatal development of the cellular and circuit properties of striatal D1 and D2 spiny projection neurons.

Key Points: Imbalances in the activity of the D1-expressing direct pathway and D2-expressing indirect pathway striatal projection neurons (SPNs) are thought to contribute to many basal ganglia disorders, including early-onset neurodevelopmental disorders such as obsessive-compulsive disorder, attention deficit hyperactivity disorder and Tourette's syndrome. This study provides the first detailed quantitative investigation of development of D1 and D2 SPNs, including their cellular properties and connectivity within neural circuits, during the first postnatal weeks. This period is highly dynamic with many properties changing, but it is possible to make three main observations: many aspects of D1 and D2 SPNs progressively mature in parallel; there are notable exceptions when they diverge; and many of the defining properties of mature striatal SPNs and circuits are already established by the first and second postnatal weeks, suggesting guidance through intrinsic developmental programmes.