Journal: bioRxiv

Article Title: Rare longevity-associated variants, including a reduced-function mutation in cGAS, identified in multigenerational long-lived families

doi: 10.64898/2025.12.04.689698

Figure Lengend Snippet: A-B ) Quantitative PCR (qPCR) analysis of downstream gene expression ( A ) and western blot analysis of phosphorylated mediators ( B ) in the canonical cGAS-STING pathway in human mesenchymal stromal cells (hMSCs) expressing the longevity-associated cGAS variant (cGAS- rs200818241), wild-type cGAS (cGAS-WT), or an empty vector control (Ctrl). C ), qPCR analysis of downstream gene expression in the canonical cGAS-STING pathway in human granulosa (KGN) cells expressing cGAS-rs200818241, cGAS-WT, or Ctrl. D ), Western blot analysis of cGAS protein levels in fibroblasts (IMR-90) and primary human astrocytes expressing cGAS-rs200818241, cGAS-WT, or Ctrl. E ), Quantification of cGAS protein levels in hMSCs, normalized to GAPDH, showing reduced levels in cells expressing cGAS-rs200818241 compared to cGAS-WT. F ), Cycloheximide (CHX) chase assay showing faster degradation of the cGAS protein in hMSCs expressing cGAS-rs200818241 compared to cGAS-WT. All experiments were performed in triplicate. Statistical significance is denoted as follows: *p < 0.05, **p < 0.01, ***p < 0.001.

Article Snippet: Human mesenchymal stem cells (hMSCs, ATCC) were cultured in Minimum Essential Medium alpha (MEM α) supplemented with 10% fetal bovine serum (FBS, Gibco), 1% penicillin/streptomycin (Gibco), and 1 ng/ml recombinant human basic fibroblast growth factor (bFGF, STEMCELL Technologies) on gelatin-coated culture dishes (Invitrogen).

Techniques: Real-time Polymerase Chain Reaction, Gene Expression, Western Blot, Expressing, Variant Assay, Plasmid Preparation, Control

Journal: bioRxiv

Article Title: Rare longevity-associated variants, including a reduced-function mutation in cGAS, identified in multigenerational long-lived families

doi: 10.64898/2025.12.04.689698

Figure Lengend Snippet: A ) Population doubling curves of cells expressing wild-type cGAS (cGAS-WT), the cGAS mutant (cGAS-rs200818241), or an empty vector control (Ctrl) in human mesenchymal stromal cells (hMSCs). Each dot indicates the cumulative population doubling per passage. cGAS-rs200818241 significantly extends cellular lifespan compared to cGAS-WT. B ) qPCR analysis of downstream genes in the canonical cGAS-STING signaling pathway and the senescence-associated p16 gene ( CDKN2A ). Expression of cGAS-rs200818241 increases both downstream genes in the canonical cGAS-STING pathway and p16 levels, while cGAS-rs200818241 reduces the expression of all these genes compared to cGAS-WT. C ), Senescence-associated β-galactosidase (SA-β-gal) staining assay showing that cGAS-WT increases the proportion of β-gal-positive cells, whereas cGAS-rs200818241 decreases the number of β-gal-positive cells compared to cGAS-WT. The left panel shows representative images of nuclei and SA-β-gal staining, the right panel presents the statistical analysis of β-gal-positive cells. All experiments were performed in triplicate. Statistical significance is denoted as follows: *p<0.05, **p < 0.01, ***p < 0.001.

Article Snippet: Human mesenchymal stem cells (hMSCs, ATCC) were cultured in Minimum Essential Medium alpha (MEM α) supplemented with 10% fetal bovine serum (FBS, Gibco), 1% penicillin/streptomycin (Gibco), and 1 ng/ml recombinant human basic fibroblast growth factor (bFGF, STEMCELL Technologies) on gelatin-coated culture dishes (Invitrogen).

Techniques: Expressing, Mutagenesis, Plasmid Preparation, Control, Staining