Biology subjects

Lindgren, J.

Publications and source records attributed to Lindgren, J..

5 recordsLinked to original sources

bioRxiv2026

Single-Cell and Tissue-Specific CRISPR Editing Analyses Unveil New Insights to Off-Targets and Translocations

CRISPR-Cas9 holds promise for treating genetic disease, but rare off-target mutations and structural variants remain as key safety concerns, especially at scales relevant to therapy. We established workflows to resolve Cas9 off-target activity in vitro at single-cell resolution and in vivo across different tissues. Using clonally expanded electroporated mouse embryos and embryonic stem cells, we reveal that individual cells exhibit unique off-target and translocation profiles, including events missed in bulk analyses. Integrating single-cell editing with chromatin accessibility, transcription, and DNA methylation measurements suggested that sequence-independent features modulate Cas9 access and cleavage, with preferential editing in regions characterized by open chromatin and lower methylation. In Cas9-inducible mouse models, editing analyses revealed organ-distinct off-target spectra, DNA repair pathway usage, indel patterns, and markedly varying translocation propensity between tissues. These findings demonstrate that off-target activity is heterogeneous across cells and context-dependent across organs, motivating sensitive single-cell analyses and organ-specific evaluation in preclinical development to more accurately assess risk and improve the safety of CRISPR-based genomic medicines.

genomics↗

bioRxiv2025

Artemis Regulates Homology-Independent Prime Editing (PRINS) for Enhanced Genomic Insertions

Nuclease-based prime editing (PEn) offers enhanced genomic insertion efficiency compared to nickase-based prime editors, but its reliance on double-strand break (DSB) repair leads to complex and often unpredictable on-target indel distributions. PRINS editing, a PEn variant utilizing springRNAs, uniquely relies on non-homologous end joining (NHEJ) for insertions, providing an insertion-only strategy ideal for functional protein tagging or serine integrase landing pad insertions, yet it suffers from inherent imprecision. Here, we identify the DNA repair factor Artemis (DCLRE1C) as a key regulator of PEn-generated indel profiles, particularly in springRNA-mediated PRINS editing. Through a targeted genetic screen, we show that the absence of Artemis significantly shifts indel distributions away from deletions and shorter truncations towards longer, functionally acceptable insertions. Our data indicates that Artemis cleaves PEn-generated 3-overhangs in a length-dependent manner, with its impact increasing for longer reverse-transcribed overhangs. This understanding reveals that regulating Artemis activity can improve insertion frequency specifically for PRINS. We develop and validate robust epigenetic (CRISPRoff) and antisense oligonucleotide (ASO) strategies to effectively silence/knockdown Artemis expression, successfully recapitulating the beneficial PRINS editing outcomes observed in Artemis-deficient cells. Leveraging these insights, we show that Artemis modulation can enhance endogenous protein tagging in cells, including challenging hiPSC-derived non-dividing cardiomyocytes. Our findings support Artemis as a key regulator of PRINS editing outcomes and present a tunable strategy to optimize insertion efficiency for diverse genomic engineering and therapeutic applications.

molecular biology↗

bioRxiv2023

Translational mapping of spatially resolved transcriptomes in human and mouse pulmonary fibrosis

Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease with poor prognosis and limited treatment options. Efforts to identify effective treatments are thwarted by limited understanding of IPF pathogenesis and poor translatability of available preclinical models. To address these limitations, we generated spatially resolved transcriptome maps of human IPF and bleomycin-induced mouse lung fibrosis. We uncovered distinct fibrotic niches in the IPF lung, characterized by aberrant alveolar epithelial cells in a microenvironment dominated by TGF{beta} signaling alongside factors such as p53 and ApoE. We also identified a clear divergence between the arrested alveolar regeneration in the IPF fibrotic niches, and the active tissue repair in the acutely fibrotic mouse lung. Our study offers in-depth insights into the IPF transcriptional landscape and proposes alveolar regeneration as a promising therapeutic strategy for IPF.

molecular biology↗

bioRxiv2022

Epigenetic modulators link mitochondrial redox homeostasis to cardiac function

Excessive production of reactive oxygen species (ROS) is characteristic of numerous diseases, but most studies in this area have not considered the impact of endogenous antioxidative defenses. Here, utilizing multi-omics, we demonstrate that in cardiomyocytes mitochondrial isocitrate dehydrogenase (IDH2) constitutes a major antioxidant defense. In both male and female mice and humans the paradoxical reduction in expression of IDH2 associated with heart failure is compensated for by an increase in the enzymes activity. We describe extensive mutual regulation of the antioxidant activities of IDH2 and NRF2 by a network involving 2-oxoglutarate and L2-hydroxyglutarate and mediated in part through unconventional hydroxymethylation of cytosine residues present in introns. Conditional targeting of ROS in a murine model of heart failure improves cardiac function. Together, these insights may explain why previous attempts to treat heart failure with antioxidants have been unsuccessful and open new approaches to personalizing and, thereby, improving such treatment. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/485908v1_ufig1.gif" ALT="Figure 1"> View larger version (66K): org.highwire.dtl.DTLVardef@1032ebeorg.highwire.dtl.DTLVardef@1f1c7bforg.highwire.dtl.DTLVardef@70f6a0org.highwire.dtl.DTLVardef@4fafbc_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIParadoxical downregulation of mitochondrial isocitrate dehydrogenase (IDH2) in response to oxidative stress leads to the discovery of a robust antioxidative defense in the heart. C_LIO_LIAn antioxidative loop involving IDH2 coordinates other antioxidative defenses, such as NRF2. C_LIO_LIThis loop produces epigenetic modifications that link oxidative stress to mitochondrial function. C_LIO_LIThe conclusion that enhancing antioxidative capacity improves cardiac function only when the endogenous capacity is insufficient opens new approaches to individualized treatment of patients with heart failure. C_LI

molecular biology↗

bioRxiv2020

A broad exome study of the genetic architecture of asthma reveals novel patient subgroups

IntroductionAsthma risk is a complex interplay between genetic susceptibility and environment. Despite many significantly-associated common variants, the contribution of rarer variants with potentially greater effect sizes has not been as extensively studied. We present an exome-based study adopting 24,576 cases and 120,530 controls to assess the contribution of rare protein-coding variants to the risk of early-onset or all-comer asthma. MethodsWe performed case-control analyses on three genetic units: variant-, gene- and pathway-level, using sequence data from the Scandinavian Asthma Genetic Study and UK Biobank participants with asthma. Cases were defined as all-comer asthma (n=24,576) and early-onset asthma (n=5,962). Controls were 120,530 UK Biobank participants without reported history of respiratory illness. ResultsVariant-level analyses identified statistically significant variants at moderate-to-common allele frequency, including protein-truncating variants in FLG and IL33. Asthma risk was significantly increased not only by individual, common FLG protein-truncating variants, but also among the collection of rare-to-private FLG protein-truncating variants (p=6.8x10-7). This signal was driven by early-onset asthma and did not correlate with circulating eosinophil levels. In contrast, a single splice variant in IL33 was significantly protective (p=8.0x10-10), while the collection of remaining IL33 protein-truncating variants showed no class effect (p=0.54). A pathway-based analysis identified that protein-truncating variants in loss-of-function intolerant genes were significantly enriched among individuals with asthma. ConclusionsAccess to the full allele frequency spectrum of protein-coding variants provides additional clarity about the potential mechanisms of action for FLG and IL33. Beyond these two significant drivers, we detected a significant enrichment of protein-truncating variants in loss-of-function intolerant genes.

genomics↗