progerin  (Addgene inc)

Journal: bioRxiv

Article Title: STING causes replication stress and nascent DNA degradation via SAMHD1

doi: 10.64898/2026.03.28.714577

Figure Lengend Snippet: ( A ) Western blot analysis of subcellular fractions from HDF treated for 6 days with doxycycline (Dox) to induce progerin expression, for 24 h with hydroxyurea (HU), or transfected with single-stranded DNA (ssDNA). Fraction markers include β-tubulin (cytoplasm), SEC61 (membrane), Lamin A (nucleus), and Histone H3 (chromatin). Note the increased presence of STING at nucleus and chromatin upon replication stress. Membranes imaged with prolonged exposure (high exposure) show a marked increase in signal intensity. ( B ) Immunofluorescence (IF) with STING antibody in HDF treated with vehicle, ssDNA, HU, Doxy (progerin), or dsDNA. ( C ) Quantification of cGAMP levels measured by ELISA in HDF treated as indicated. ( D ) IF with STING antibody and quantification of percentage of cells showing STING localization to the perinuclear compartment (PNC) upon different treatments. ( E ) IF with antibody recognizing phosphorylated STING on Ser366 and quantification of percentage of cells positive for S366 p-STING. ( F ) Immunoblot analysis of STING, GFP-progerin, and markers of activation of the canonical cGAS-STING pathway ( S366 p-STING, S386 p-IRF3, and S172 p-TBK1) following treatments. ( G ) Immunoblot analysis of STING pathway components and ISG proteins (STAT1, S727 p-STAT1, RIG-I, and ISG15) after indicated treatments.

Article Snippet: Primary antibodies used were β-tubulin (1:5000- Origene-AP31823PU-N), GAPDH (1:1000- Cell Signaling-2118), Lamina A (1:3000- Abcam-1791), Progerin (1:1000 Santa Cruz-81511), ISG15 (1:1000 Santa Cruz-166755), STING (1:1000-Cell Signaling-13647), S366 p-STING (1:1000- Cell Signaling- 50907), RIG-I (1:1000-Cell Signaling-3743S), S33-p-RPA (1:1000 Bethyl- PLA0070), SAMHD1 (1:1000- Cell Singaling-49158), λH2AX (1:1000- Cell Signaling- 2577).

Techniques: Western Blot, Expressing, Transfection, Membrane, Immunofluorescence, Enzyme-linked Immunosorbent Assay, Activation Assay

Journal: bioRxiv

Article Title: STING causes replication stress and nascent DNA degradation via SAMHD1

doi: 10.64898/2026.03.28.714577

Figure Lengend Snippet: ( A ) U2OS tumor cells were generated that express progerin, STING, or both, using empty vectors (EV) as controls. Immunoblots performed to assess replication stress marker 33 p-RPA, and activation of an IFN response (ISG15, RIG-I). ( B ) DNA fiber assays to monitor total tract length in U2OS +/- progerin and +/- STING. ( C ) Fork symmetry (ratio of the sisters CldU tracts from bidirectional forks) in same conditions as in (B). ( D ) DNA fiber analysis to monitor total tract length in U2OS +/- progerin and +/- STING with or without dNTP supplementation. ( E ) Fork symmetry in the same samples as in (D). ( F ) Immunoblot showing STING expression and SAMHD1 knockdown in control and progerin-expressing U2OS cells. ( G ) DNA fiber analysis to monitor total tract length in U2OS +/- progerin and +/- STING with or without SAMHD1 depletion. ( H ) Fork symmetry in the same samples as in (G). ( I ) Subcellular fractionation of U2OS +/- progerin and +/- STING, probed for STING and SAMHD1. H3 and SEC61 were used as nuclear and ER markers, respectively. For all quantifications, each point represents a single replication fork; n = 3 biologically independent experiments.

Article Snippet: Primary antibodies used were β-tubulin (1:5000- Origene-AP31823PU-N), GAPDH (1:1000- Cell Signaling-2118), Lamina A (1:3000- Abcam-1791), Progerin (1:1000 Santa Cruz-81511), ISG15 (1:1000 Santa Cruz-166755), STING (1:1000-Cell Signaling-13647), S366 p-STING (1:1000- Cell Signaling- 50907), RIG-I (1:1000-Cell Signaling-3743S), S33-p-RPA (1:1000 Bethyl- PLA0070), SAMHD1 (1:1000- Cell Singaling-49158), λH2AX (1:1000- Cell Signaling- 2577).

Techniques: Generated, Western Blot, Marker, Activation Assay, Expressing, Knockdown, Control, Fractionation

Journal: bioRxiv

Article Title: Senescence-inhibitory Δ133p53α counteracts accelerated ageing and mortality

doi: 10.64898/2025.12.31.697195

Figure Lengend Snippet: Tissue sections of the aorta and skin were prepared from Group-1, Group-3, and wild-type (WT) mice at 9-10 months of age (n = 5 each; open circles, females; closed circles, males). a, Restoration of vascular smooth muscle cells (VSMCs) in the aorta by Δ133p53α. Representative images of the serial sections stained with hematoxylin and eosin (H&E, top) and Verhoeff–Van Gieson (VVG, bottom) are shown (scale bars, 50 µm). The tunica media was identified by black-stained elastic fibers. Cell numbers in the tunica media (per mm 3 ) are presented as mean ± s.d. (n = 5). b, Restoration of dermal thickness in the skin by Δ133p53α. Thickness of the dermis (µm) for each mouse was calculated as the average of measurements taken at five randomly selected, approximately evenly spaced sites. Scale bars, 200 µm. Data are presented as mean ± s.d. (n = 5). Epi, epidermis; D, dermis; dWAT, dermal white adipose tissue. c, Immunofluorescence (IF) staining of Sox9 in the hair follicles. Representative images of Sox9 (green) and DAPI (blue) are shown (scale bars, 50 µm). All five mice in each group showed highly reproducible results. d, Sox9 mRNA expression analyzed by qRT-PCR in the skin of Group-1, Group-3 and wild-type mice. Data are presented as mean ± s.d. (n = 5, each with technical triplicate). e, Sox9 mRNA expression in MEFs. CAG-133 LSL/+ ; Cre Tg/+ MEFs were with or without retroviral progerin expression, and with or without 4-OHT-induced Δ133p53α expression. f, SOX9 mRNA expression in HGPS patient-derived fibroblasts. Two fibroblast strains, AG11513 and HGADFN271, were transduced with Δ133p53α-expressing (+) or control (-) lentiviral vectors. Data are presented as mean ± s.d. (n = 3, technical triplicate) ( e,f ). P values were calculated by Welch’s t -test ( a,b,d,e,f ).

Article Snippet: The retroviral vector for expression of progerin (pBABE-puro-GFP-progerin) was obtained from Addgene (Plasmid #17663).

Techniques: Staining, Immunofluorescence, Expressing, Quantitative RT-PCR, Retroviral, Derivative Assay, Transduction, Control