Journal: PNAS Nexus

Article Title: Ancient and animal-specific regulatory modes of EWS::FLI1 revealed by a minimal yeast model

doi: 10.1093/pnasnexus/pgag329

Figure Lengend Snippet: Introduction of human GGAA microsatellites into the yeast genome induces gene silencing. A) Constructs carrying eGFP driven by the minimal pCYC1 promoter with upstream 80-nt random or 10×/20×/40 × (GGAA) inserts. B) Confocal images of W303-1A isogenic derivatives carrying the constructs shown in (A): GFP fluorescence (right) and bright field (left); scale bar, 10 μm. C, D) Quantification by flow cytometry ( n = 10,000 cells per sample). C) Distribution of cell counts as a function of Comp-GFP. D) Mean eGFP fluorescence.

Article Snippet: GFP fluorescence was assessed by SP8 confocal microscopy and by spectral flow cytometry (Cytek Aurora; 10,000 cells/sample).

Techniques: Yeast, Construct, Confocal, Fluorescence, Flow Cytometry

Journal: PNAS Nexus

Article Title: Ancient and animal-specific regulatory modes of EWS::FLI1 revealed by a minimal yeast model

doi: 10.1093/pnasnexus/pgag329

Figure Lengend Snippet: Transcriptional activation by EWS::FLI1 through recognition of GGAA microsatellites in yeast. A) W303-1A derivatives carrying pCYC1 (top), 20 × (GGAA) (middle), or 40 × (GGAA) (bottom) reporters transformed with empty pYES2, EWS::FLI1, or EWS::FLI1 RRLL . Left, representative confocal images (bright-field, top; GFP fluorescence, bottom); middle, flow-cytometry histograms of cell counts as a function of Comp-GFP; right, mean GFP fluorescence. B) Time-course of 20×(GGAA)_ pCYC1 _eGFP cells transformed with pYES2 carrying EWS::FLI1 RRLL (top) or EWS::FLI1 (bottom). Samples collected at the indicated times were immunoblotted with anti-EWSR1 and anti-GFP monoclonal antibodies; arrowheads indicate EWS::FLI1 and GFP. Ponceau staining is shown as a loading and transfer control. Equivalent results were obtained in two further biological replicates. C) Relative EWS::FLI1 and GFP levels, normalized to the mean EWS::FLI1 signal at 6 h of induction (100%). Error bars, SD. * P < 0.05; ns, not significant; two-way ANOVA.

Article Snippet: GFP fluorescence was assessed by SP8 confocal microscopy and by spectral flow cytometry (Cytek Aurora; 10,000 cells/sample).

Techniques: Activation Assay, Yeast, Transformation Assay, Confocal, Fluorescence, Flow Cytometry, Bioprocessing, Staining, Control

Journal: PLOS Pathogens

Article Title: Elevated Plasmodium falciparum sexual conversion in HbAC and HbAS red blood cells in naturally infected malaria patients

doi: 10.1371/journal.ppat.1014542

Figure Lengend Snippet: (A) Experimental design of in vitro SC assays ( iv SCAs) using the NF54 -gexp02-Tom P. falciparum line. Tightly synchronized schizonts (Generation -1) were allowed to reinvade red blood cells (RBCs) of different HBB genotypes, and ring stages (0-5 hpi) were cultured under non-inducing (+Choline) and inducing (-Choline) conditions. Parasite development was assessed by flow cytometry at 35-40 hpi (Generation 0) and 25-30 hpi/35-40 hpi (Generation 1). SCRs were measured by flow cytometry at 25-30 hpi and 35-40 hpi (Generation 1). SCRs by HBB genotype at 35-40 hpi (Generation 1) in (B) +Choline cultures and (C) in -Choline cultures. (D) Proportion of asexual and gametocyte stages and total parasitemia by HBB genotype at 35-40 hpi (Generation 1) in +Choline cultures. (E) Reticulocytes percentage by HBB genotype. Statistical tests: B, C and E, Beta-binomial GLMs with Bonferroni correction for pairwise comparisons. Significant p -values are shown as: p < 0.05*, p < 0.01**, p < 0.001***; non-significant p -values as: ns. Abbreviations: CTRL, control sample; GLMs, Generalized linear models; Hb, hemoglobin; HBB, human hemoglobin beta; Hpi, hours post-invasion; SC, sexual conversion; SCR, sexual conversion rates. Panel A was created in BioRender. Yasmina, D. (2026) https://BioRender.com/ew8vbur .

Article Snippet: Samples for flow cytometry analysis (using a CytoFLEX LX flow cytometer, Beckman Coulter) were prepared by resuspending parasite cultures (1:3) in PBS containing 1% BSA, followed by staining with Hoechst 33342 (1:500, BD Pharmingen) in a final volume of 100 μl per well.

Techniques: In Vitro, Cell Culture, Parasite, Flow Cytometry, Control

Journal: PLOS Pathogens

Article Title: Elevated Plasmodium falciparum sexual conversion in HbAC and HbAS red blood cells in naturally infected malaria patients

doi: 10.1371/journal.ppat.1014542

Figure Lengend Snippet: (A) Microscopy-based quantification of parasite stages (rings, trophozoites and schizonts) expressed as percentages of a total of 100 counted parasites per sample. Y-axis: percentage of parasite stages (%); X-axis: HBB genotype groups (HbAA n = 1, HbAC n = 1 and HbAS n = 2, and HbSS n = 2 and HbSC n = 2 donors from a malaria-endemic area in Nanoro, Burkina Faso. Timepoints shown are where progression toward late stages ( i.e., schizont stages) is most clearly observed, namely 35–40 hpi (Generation 0; left graph and Generation 1; right graph). A shift towards earlier parasite stages indicates developmental delay.(B) Developmental progression of individual parasite cultures across timepoints 35–40 hpi (Generation 0), and 25–30 hpi and 35–40 hpi (Generation 1), shown as the proportion of late stages (schizonts) among total infected RBCs (iRBCs) measured by flow cytometry. Data are stratified by HBB genotype: HbAA ( n = 5), HbAC ( n = 5),HbAS ( n = 4), and HbSC ( n = 1) donors from a malaria-endemic area in Nanoro, Burkina Faso; and HbSS ( n = 12), HbSC ( n = 1) and n = 6 control HbAA donors [HbAA CTRL] from a non-endemic area. (C) Representative flow cytometry plots showing the gating strategy for the detection of late parasite stages. Left: total RBCs gated. Right: within the total RBCs gate in left plot, uninfected RBCs (uRBCs) and iRBCs are identified, with iRBCs further gated into early and late parasite stages by Hoechst 33342. Abbreviations: CTRL, control sample; FSC-A, Forward Scatter-Area; G0, Generation 0; G1, Generation 1; GLMs, Generalized linear models; Hb, hemoglobin; HBB, human hemoglobin beta; Hpi, hours post-invasion; RBCs, red blood cells; SSC-A, Side Scatter-Area; Timepoint abbreviations are specified in the figure legend.

Article Snippet: Samples for flow cytometry analysis (using a CytoFLEX LX flow cytometer, Beckman Coulter) were prepared by resuspending parasite cultures (1:3) in PBS containing 1% BSA, followed by staining with Hoechst 33342 (1:500, BD Pharmingen) in a final volume of 100 μl per well.

Techniques: Microscopy, Parasite, Infection, Flow Cytometry, Control