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In primary cell types, intracellular deoxynucleotide triphosphate (dNTP) levels are tightly regulated in a cell cycle-dependent manner. We report that prime editing efficiency is increased by mutations that improve the enzymatic properties of Moloney murine leukemia virus reverse transcriptase and treatments that increase intracellular dNTP levels. In combination, these modifications produce substantial increases in precise editing rates., (© 2024. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Identifying the early islet cellular processes of autoimmune type 1 diabetes (T1D) in humans is challenging given the absence of symptoms during this period and the inaccessibility of the pancreas for sampling. In this article, we study temporal events in pancreatic islets in LEW.1WR1 rats, in which autoimmune diabetes can be induced with virus infection, by performing transcriptional analysis of islets harvested during the prediabetic period. Single-cell RNA-sequencing and differential expression analyses of islets from prediabetic rats reveal subsets of β- and α-cells under stress as evidenced by heightened expression, over time, of a transcriptional signature characterized by interferon-stimulated genes, chemokines including Cxcl10, major histocompatibility class I, and genes for the ubiquitin-proteasome system. Mononuclear phagocytes show increased expression of inflammatory markers. RNA-in situ hybridization of rat pancreatic tissue defines the spatial distribution of Cxcl10+ β- and α-cells and their association with CD8+ T cell infiltration, a hallmark of insulitis and islet destruction. Our studies define early islet transcriptional events during immune cell recruitment to islets and reveal spatial associations between stressed β- and α-cells and immune cells. Insights into such early processes can assist in the development of therapeutic and prevention strategies for T1D., (© 2023 by the American Diabetes Association.)

New variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continue to emerge, causing surges, breakthrough infections, and devastating losses-underscoring the importance of identifying SARS-CoV-2 antivirals. A simple, accessible human cell culture model permissive to SARS-CoV-2 variants is critical for identifying and assessing antivirals in a high-throughput manner. Although human alveolar A549 cells are a valuable model for studying respiratory virus infections, they lack two essential host factors for SARS-CoV-2 infection: angiotensin-converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2). SARS-CoV-2 uses the ACE2 receptor for viral entry and TMPRSS2 to prime the SARS-CoV-2 spike protein, both of which are negligibly expressed in A549 cells. Here, we report the generation of a suitable human cell line for SARS-CoV-2 studies by transducing human ACE2 and TMPRSS2 into A549 cells. We show that subclones highly expressing ACE2 and TMPRSS2 ("ACE2plus" and the subclone "ACE2plusC3") are susceptible to infection with SARS-CoV-2, including the delta and omicron variants. These subclones express more ACE2 and TMPRSS2 transcripts than existing commercial A549 cells engineered to express ACE2 and TMPRSS2. Additionally, the antiviral drugs EIDD-1931, remdesivir, nirmatrelvir, and nelfinavir strongly inhibit SARS-CoV-2 variants in our infection model. Our data show that ACE2plusC3 cells are highly permissive to SARS-CoV-2 infection and can be used to identify anti-SARS-CoV-2 drugs.