Biology subjects

Lazarchuk, P.

Publications and source records attributed to Lazarchuk, P..

3 recordsLinked to original sources

bioRxiv2025

Cellular heterogeneity and therapeutic response profiling of human IDH+ glioma stem cell cultures

Summary/AbstractGlioblastoma stem cell (GSC) cultures are initiated from glioblastoma (GBM) surgical resection tissue. They can capture and propagate key GBM primary tumor molecular and cellular features. We have deeply characterized four isocitrate dehydrogenase (IDH)-expressing (or IDH+) GSC cultures from unrelated adults to serve as cellular models for the majority of adult primary GBM. We demonstrate that GSC cultures can be continuously propagated in defined, serum-free media and 5% oxygen without requiring specialized growth substrates; have well-defined genomic and mtDNA variants and gene/protein expression profiles; and highly reproducible dose-survival curves when treated with the GBM standard-of-care therapies of ionizing radiation (IR) and temozolomide (TMZ). We also illustrate how expressed lentiviral barcodes, mtDNA variants and single cell gene expression profiling can be used to define and track cellular heterogeneity over 40 days after IR treatment. These well-characterized IDH+ GSC cultures can support many high throughput in vitro assay formats, including xenograft, organoid and other GBM disease modeling protocols. They should prove a useful resource to better understand GBM biology, and to identify new and more effective GBM therapies and treatment regimens. HighlightsO_LIGlioblastoma (GBM)-derived IDH-expressing Glioma Stem Cell (GSC) cultures can capture and propagate GBM genomic variants, gene and protein expression programs. C_LIO_LIMitochondrial DNA (mtDNA) variants identified by high accuracy Duplex DNA sequencing are abundant and sub-clonally organized in GSC cultures. C_LIO_LIGSC cultures have highly reproducible dose-survival curves for ionizing radiation and temozolomide, the GBM standard-of-care therapies. C_LIO_LIGSC culture cellular heterogeneity can be captured, characterized and tracked by using expressed lentiviral barcodes, mtDNA variants and scRNA sequencing. C_LI eTOC blurbIDH-expressing glioma stem cell cultures (GSCs) are experimentally tractable and versatile glioblastoma (GBM) cellular disease models. Deeply characterized GSC cultures can enable new work to understand GBM biology, and help identify new and more effective GBM therapies and treatment regimens. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=113 SRC="FIGDIR/small/667532v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@82aceeorg.highwire.dtl.DTLVardef@1277c18org.highwire.dtl.DTLVardef@144188eorg.highwire.dtl.DTLVardef@1a84bb3_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2024

Werner syndrome RECQ helicase participates in and directs maintenance of the protein complexes of constitutive heterochromatin in proliferating human cells

The WRN RECQ helicase is responsible for the Werner syndrome of premature aging and cancer predisposition. Substantial progress has been made in delineating WRN functions in multiple aspects of DNA metabolism, including DNA replication, repair, transcription, and telomere maintenance. Nevertheless, a complete mechanistic understanding of how loss of WRN accelerates aging in humans has not been achieved yet. Here we show that WRN is involved in the maintenance of constitutive heterochromatin, CH, in proliferating, immortalized human fibroblasts. WRN is found within a complex with histone deacetylase 2, HDAC2, and WRN/HDAC2 association is mediated by heterochromatin protein alpha, HP1. WRN deficiency derepresses SATII pericentromeric satellite repeats and reduces a subset of protein-protein interactions that participate in the organization of CH in the nucleus. In particular, WRN deficiency reduces the complexes involving Lamin B1 and Lamin B receptor, LBR. Both mRNA level and subcellular distribution of LBR are affected by WRN deficiency, and the latter phenotype does not require WRN catalytic activities. At the mRNA level, WRN supports complete maturation of the LBR mRNA. All signs of heterochromatin disruption seen in WRN-deficient proliferating fibroblasts are also observed in WRN-proficient fibroblasts undergoing replicative or oncogene-induced senescence, and WRN complexes with HP1 and HDAC2 are also markedly downregulated in these senescing cells. The data suggest that WRN loss affects heterochromatin independently of the senescence program but can mimic aspects of it and thus sensitize cells to triggers of senescence.

molecular biology↗

bioRxiv2021

Innate immunity mediator STING modulates nascent DNA metabolism at stalled forks in human cells

The cGAS/STING pathway, part of the innate immune response to foreign DNA, is known to be activated by cells own DNA arising from the processing of the genome, including the excision of nascent DNA at arrested replication forks. We found STING activation to affect nascent DNA processing, suggesting a novel, unexpected feedback connection between the two events. Depletion of STING suppressed and re-expression of the protein in STING-deficient cells upregulated degradation of nascent DNA. Fork arrest was accompanied by the STING pathway activation, and a STING mutant that does not activate the pathway failed to upregulate nascent strand degradation. Consistent with this, cells expressing the STING mutant had a reduced level of RPA on parental and nascent DNA of arrested forks as well as a reduced CHK1 activation compared to the cells with wild type STING. Together our findings reveal a novel connection between replication stress and innate immunity.

molecular biology↗