Journal: bioRxiv

Article Title: Oncogenic virus hijacks SOX18 pioneer function to enhance viral persistence

doi: 10.1101/2025.06.28.662102

Figure Lengend Snippet: A-C. HeLa cells expressing SOX18wt, mutants C240X (dominant-negative transactivation deficient) or HMGdel (DNA-binding deficient), or mCherry as a control, and thereafter infected with rKSHV.219 for 72h (KSHV-HeLa). A. IF images of the SOX18wt and mutants expressing cells labeled with anti-SOX18 antibody and a schematic of the constructs. B. Immunoblotting with anti-SOX18 antibody using β-actin as a loading control for normalization. C. RT-qPCR for the indicated viral genes in KSHV-HeLa. D. LECs infected with rKSHV.219 (KLECs) for 72 hours and treated with Sm4 or DMSO control for 24h and relative mRNA measured for indicated viral transcripts. Statistical significance was determined by one-way ANOVA with Dunnett correction for multiple comparisons; ns = non-significant. E-F. Uninfected LECs and KLECs 72h p.i. treated with DMSO or Sm4 for another 72h and E) labeled with anti-ARID1A and -BRG1 antibodies, nuclei were counterstained with Hoechst (33342), scale bar is 10µm, and F) immunoblotted for the indicated proteins and quantified as in B.

Article Snippet: Primary human dermal lymphatic endothelial cells LEC (Promocell; C-12216) were maintained in Microvascular MV-2 (Promocell; C-22121) medium supplemented with 5% fetal bovine serum, basic fibroblast growth factor, insulin-like growth factor 3, epidermal growth factor, gentamicin sulfate/amphotericin, ascorbic acid, and hydrocortisone; VEGF was not added.

Techniques: Expressing, Dominant Negative Mutation, Binding Assay, Control, Infection, Labeling, Construct, Western Blot, Quantitative RT-PCR

Journal: bioRxiv

Article Title: Oncogenic virus hijacks SOX18 pioneer function to enhance viral persistence

doi: 10.1101/2025.06.28.662102

Figure Lengend Snippet: A-C. A schematic of the inhibitor mode of action is shown in the top panels. CTG viability assay of uninfected LECs (LEC) or LECs infected with rKSHV.219 (KLEC) for 72h and treated with the indicated, increasing concentrations of BRG1 inhibitors A) ACBI1, B) FHT-1015 and C) PFI-3 (n=3), arrows indicate the selected concentration for following inhibitor assays. D. LECs and KLECs were treated with ACBI1, FHT-1015 and PFI-3 and treated with EdU for 4h before subjecting to EdU Click-It, imaged with Opera Phenix 20x and quantified from (n=6 independent replicates, and from each n=100 nuclei). Statistical significance was determined by one-way ANOVA with Dunnett correction for multiple comparisons, ns = non-significant. E-F. LECs infected with ΔORF50 and treated with Sm4, FHT-1015, or DMSO for 72h and processed for ATAC-seq. E. Peaks and heatmaps of the top 1000 genomic regions with reduced overall accessibility (dark blue maps) showing shared (dark blue line), unique to Sm4 (turquoise) and unique to FHT-1015 (purple) loss sites. F. Pearson’s analysis of the replicate (n = 3) samples. ns = non-significant.

Article Snippet: Primary human dermal lymphatic endothelial cells LEC (Promocell; C-12216) were maintained in Microvascular MV-2 (Promocell; C-22121) medium supplemented with 5% fetal bovine serum, basic fibroblast growth factor, insulin-like growth factor 3, epidermal growth factor, gentamicin sulfate/amphotericin, ascorbic acid, and hydrocortisone; VEGF was not added.

Techniques: Viability Assay, Infection, Concentration Assay

Journal: PLOS Pathogens

Article Title: BiP/GRP78 is a pro-viral factor for diverse dsDNA viruses that promotes the survival and proliferation of cells upon KSHV infection

doi: 10.1371/journal.ppat.1012660

Figure Lengend Snippet: (A-B) HA15 treatment differentially reduces cell numbers compared to cell viability in TREx-BCBL-1 cells. Latent TREx-BCBL-1 cells were treated with increasing concentrations of HA15 (0–50 μM) for 72h. The total number of viable cells (A) and the percent of dead TREx-BCBL-1 cells (B) were determined by automated cell counting following trypan blue staining. (C-D) HA15 treatment does not cause cytostasis nor cytotoxicity in primary B cells. Primary Peripheral B-cells were treated with increasing concentrations of HA15 (0–50 μM) for 72h. The total number of viable cells (C) and the percent of live cells (D) were determined as described in (A-B). N = 3 independent biological replicates. Values are average ±SEM. (E-G) Primary Lymphatic endothelial cells were infected with KSHV.219 and selected with puromycin for 7–14 days. (A) Whole-cell lysates from uninfected (LEC) or infected (KLECs) were collected and analyzed by immunoblot. Actin: loading control. (B-C) LECs and KLECs were treated with HA15 (10 μM) for 72h. Cell viability was evaluated by ATP quantification using CellTiter-Glo (B) and microscopy at 0h and 72h post-treatment (C.) .

Article Snippet: Primary Dermal Lymphatic endothelial cells (LECs) from PromoCell (C-12217) were maintained in EBM-2 media (Lonza 00190860) supplemented with the EGM-2 MV bullet kit (CC-4147) at 37°C with a 5% CO2 atmosphere.

Techniques: Cell Counting, Staining, Infection, Western Blot, Control, Microscopy

Journal: The EMBO Journal

Article Title: Transcription factor FOXP2 is a flow‐induced regulator of collecting lymphatic vessels

doi: 10.15252/embj.2020107192

Figure Lengend Snippet: Isolation of dermal EC subtypes from the ear skin of 5‐week‐old Prox1‐GFP mice using multicolor flow cytometry. Whole‐mount immunofluorescence for LYVE1 (on the left) defines three Prox1‐GFP + lymphatic vessels subtypes that are sorted using the indicated gating scheme (on the right): LYVE1 low/− collecting vessel (green arrow, col); LYVE1 intermed pre‐collecting vessel (gray arrow, pre‐col); LYVE1 high lymphatic capillary (magenta arrow, cap). Top 100 genes upregulated in collecting in comparison to lymphatic capillary LECs. Expression in BECs is shown for comparison. Heat map color coding shows log 2 fold change. Red box indicates LEC‐specific/enriched collecting vessel signature genes that include Foxp2 (red arrow). Whole‐mount immunofluorescence of ear skin, mesentery, and flank, showing nuclear FOXP2 staining in collecting lymphatic vessels (arrows, col), but not in LYVE1 + lymphatic capillaries (cap) or blood vessels (bv). qRT–PCR analysis of Foxp1–4 in murine ECs freshly isolated from P11 mesentery ( n = 6 mice, individual data points shown). Data are presented as mean relative expression (normalized to Gapdh ) ± SD. Transcript levels for each transcript are presented relative to levels in LECs. P , Student's t ‐test. nd, not detected. Data information: Scale bar: 75 µm (A), 50 µm (C). Source data are available online for this figure.

Article Snippet: Human primary dermal LECs (HDLECs isolated from juvenile foreskin, cat no. C‐12216) were obtained from PromoCell.

Techniques: Isolation, Flow Cytometry, Immunofluorescence, Comparison, Expressing, Staining, Quantitative RT-PCR

Journal: The EMBO Journal

Article Title: Transcription factor FOXP2 is a flow‐induced regulator of collecting lymphatic vessels

doi: 10.15252/embj.2020107192

Figure Lengend Snippet: A Whole‐mount immunofluorescence of embryonic mesenteries of the indicated developmental stages. Note induction of FOXP2 expression at E17. B Whole‐mount immunofluorescence of E18 mesenteries fixed immediately after dissection (left panel) or after 24 h of ex vivo culture (right panel). Note loss of patterning of PROX1 high valve LECs and downregulation of FOXP2 expression in flow‐abrogated vessels after ex vivo culture. C, D Immunofluorescence (C) and qRT–PCR analysis (D) of HDLECs grown under static conditions or exposed to OSS for 48 h ( n = 3 independent experiments). Data are presented as mean ± SD. P , Student's t ‐test. Data information: Scale bar: 100 µm (A, B), 50 μm (C). Source data are available online for this figure.

Article Snippet: Human primary dermal LECs (HDLECs isolated from juvenile foreskin, cat no. C‐12216) were obtained from PromoCell.

Techniques: Immunofluorescence, Expressing, Dissection, Ex Vivo, Quantitative RT-PCR

Journal: The EMBO Journal

Article Title: Transcription factor FOXP2 is a flow‐induced regulator of collecting lymphatic vessels

doi: 10.15252/embj.2020107192

Figure Lengend Snippet: qRT–PCR analysis of GATA2 expression in control (siCTRL) and FOXP2 or FOXC2 siRNA‐treated HDLECs ( n = 3 independent experiments). Left: NFATc1 immunofluorescence in control (siCTRL) and FOXP2 or FOXC2 siRNA‐transfected HDLECs under basal conditions (untreated) or after 30‐min stimulation with VEGF‐C (50 ng/ml) in the absence or presence of the calcineurin inhibitor cyclosporin A (CsA). Note nuclear translocation of NFATc1 upon VEGF‐C stimulation, which is abrogated in cells treated with FOXP2 or FOXC2 siRNA, or CsA. Right: Quantification of nuclear NFATc1 in CTRL and FOXP2 siRNA‐treated LECs after VEGF‐C stimulation ( n = 7–9 images from two independent experiments). Data information: In (A, B), data are presented as mean ± SD. P , one‐way ANOVA (A), or Student's t ‐test (B). Scale bar: 50 µm (B). Source data are available online for this figure.

Article Snippet: Human primary dermal LECs (HDLECs isolated from juvenile foreskin, cat no. C‐12216) were obtained from PromoCell.

Techniques: Quantitative RT-PCR, Expressing, Control, Immunofluorescence, Transfection, Translocation Assay