Biology subjects

Krach, F.

Publications and source records attributed to Krach, F..

4 recordsLinked to original sources

bioRxiv2025

Deciphering Brain Organoids Heterogeneity by Identifying Key Quality Determinants

Brain organoids derived from human pluripotent stem cells (hPSCs) hold immense potential for modeling neurodevelopmental processes and disorders. However, their experimental variability and undefined organoid selection criteria for analysis hinder reproducibility. As part of the Bavarian ForInter consortium, we generated 72 brain organoids from distinct hPSC lines. We conducted a comprehensive analysis of their morphological and cellular characteristics at an early stage of their development. In our assessment, the Feret diameter emerged as a reliable, single parameter that characterizes brain organoid quality. Transcriptomic analysis further confirmed the reliability of this marker and identified a negative impact of mesenchymal cells on the abundance of organoid formation. High-quality organoids consistently displayed a lower mesenchymal cell presence. These findings offer a framework for enhancing brain organoid standardization and reproducibility, underscoring the need for morphological quality controls and the consideration of mesenchymal cell influence on organoid-based modeling. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=186 HEIGHT=200 SRC="FIGDIR/small/632763v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@11c214org.highwire.dtl.DTLVardef@1dcb643org.highwire.dtl.DTLVardef@1420141org.highwire.dtl.DTLVardef@6074e6_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2023

RNA splicing modulator induces peripheral neuropathy with increased neurofilament light chain levels via p53 signaling

RNA splicing modulators are a new class of small molecules with the potential to modify the expression levels of proteins. A recent clinical trial investigating the splicing modulator branaplam for Huntingtons disease to lower huntingtin levels was terminated due to the development of peripheral neuropathy. Here, we describe how branaplam leads to this adverse effect. On a cellular level, branaplam disrupts neurite integrity reflected by elevated neurofilament light chain levels in human induced pluripotent stem cell (iPSC)-derived motor neurons (iPSC-MN). Branaplam does not target neuropathy-associated genes. However, transcription factor binding site enrichment analysis indicates p53 activation. P53 activation upon branaplam treatment in iPSC-MN is linked to increased nucleolar stress, thereby enhanced expression of the neurotoxic p53-target gene BBC3. These findings illustrate that RNA splicing modulators may have clinically relevant off-target effects, implying the necessity of comprehensive pre-screening in human models prior to executing clinical trials. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=188 SRC="FIGDIR/small/551640v1_ufig1.gif" ALT="Figure 1"> View larger version (47K): org.highwire.dtl.DTLVardef@9b2cbdorg.highwire.dtl.DTLVardef@1964746org.highwire.dtl.DTLVardef@121612corg.highwire.dtl.DTLVardef@72c6a6_HPS_FORMAT_FIGEXP M_FIG C_FIG One Sentence SummaryPredicting side effects of RNA splicing modulator branaplam leading to neurotoxicity via nucleolar stress, p53 activation, and axonal degeneration.

pharmacology and toxicology↗

bioRxiv2017

Reversal of molecular pathology by RNA-targeting Cas9 in a myotonic dystrophy mouse model

The dominantly inherited, multi-systemic disease myotonic dystrophy type I (DM1) is caused by triplet repeat CTG expansions in the DMPK gene and is the most common form of adult-onset muscular dystrophy. Elimination of the toxic, repetitive CUG RNA constitutes a therapeutic for this disease. We report an RNA-targeting Cas9 (RCas9) system that supports efficient reversal of DM1 phenotypes via delivery to adult poly(CUG) DM1 mouse muscle using adeno-associated virus (AAV). We observe elimination of CUG RNA, restoration of CUG foci-associated Mbnl1 protein to wild-type subcellular localization, correction of DM1-type alternative splicing patterns in candidate genes including the voltage-gated chloride channel 1 (Clcn1) responsible for characteristic myotonia, recovery of Clcn1 staining, and reduction in centralized myonuclei. Our results establish RCas9 as a potential long-term in vivo therapeutic for DM1.\n\nOne Sentence SummaryA repurposed CRISPR system termed RNA-targeting Cas9 reverses the molecular pathology associated with the most common type of adult onset muscular dystrophy in adult mouse muscle.

molecular biology↗

bioRxiv2017

Proximity labeling reveals an extensive steady-state stress granule interactome and insights to neurodegeneration

Stress granules (SGs) are transient ribonucleoprotein (RNP) aggregates that form in response to proteotoxic stress. Although SGs are distinct from aggregates observed in neurodegenerative disorders, they share protein components. We used APEX-mediated proximity labeling combined with quantitative mass spectrometry and high-throughput imaging to identify >100 previously unknown SG proteins in human cells, about 10% of which localize to SGs in a cell type- or stress type-dependent manner. Supporting a link between SG proteins and neurodegeneration, we demonstrate aberrant SG composition and subcellular distribution in iPSC-derived motor neurons from ALS patients, and identify several known and previously unidentified SG proteins that modify toxicity of mutant FUS and TDP-43 overexpression in Drosophila. We show that even in an unstressed steady-state, SG proteins form a densely-connected protein interaction network (PIN) and propose a model in which existing RNPs coalesce rapidly into microscopically visible granules that can act as gateways to pathological protein aggregation.\n\nHighlights O_LIAPEX proximity labeling of dynamic RNP granules identifies over 100 novel SG proteins\nC_LIO_LISG proteins form a densely-connected protein interaction network in unstressed cells\nC_LIO_LISystematic immunofluorescence analysis reveals stress- and cell type-specific SG composition\nC_LIO_LIALS motor neurons contain SGs with distinct content and subcellular distribution\nC_LI

cell biology↗