Journal: Nature Microbiology

Article Title: Specialized RNA decay fine-tunes monogenic antigen expression in Trypanosoma brucei

doi: 10.1038/s41564-026-02289-4

Figure Lengend Snippet: a , b , Fluorescence microscopy analysis of ESB2 12myc localization following ESB3 and ESAP1 RNAi ( a , b ), 6myc ESB3 localization following ESB2 and ESAP1 RNAi ( b ) and 6myc ESAP1 localization following ESB2 and ESB3 RNAi ( b ). c , ESB2, ESB3 and ESAP1 protein levels following VEX2, ESB2, ESB3 and ESAP1 RNAi. The values were derived from protein blotting analysis of three biological replicates per cell line and are represented in a floating bar graph in which the box spans between minimum and maximum values, the centre line is the median, and all datapoints are depicted. Two-tailed unpaired t -tests were applied; NS, non-significant; ** P < 0.01. d – i , Fluorescence microscopy analysis of Pol-I localization following VEX2, ESB1, ESB2, ESB3 and ESAP1 knockdown ( d , e ); ESB1 12myc , ESB2 12myc , 6myc ESB3 and 6myc ESAP1 localization following VEX2 knockdown ( f , g ); 6myc VEX2 localization following ESB2, ESB3 and ESAP1 knockdown ( h , i ). The graphs in b , e , g and i depict mean values of two biological replicates; >100 G1 cells per condition were analysed; all analyses were performed at 24 h post-induction. j , Heatmap summarizing the microscopy analyses; C1, clone 1; C2, clone 2. k , Diagram depicting the complex network of co-dependencies at the ESB and interface with the SLAB. The direction of the arrows indicates that the protein upstream is required for the localization of the protein downstream to its corresponding nuclear compartment. l , Fluorescence microscopy colocalization analysis of GFP-myc ESB2 and 6HA ESB3. The images in a , d (left), h and l were acquired using a Zeiss AxioObserver, whereas the images in d (right) and f were acquired using a Zeiss LSM980 Airyscan 2. All correspond to 3D projections by brightest intensity of 0.1-μm stacks. DNA was stained with DAPI (cyan or grey). Scale bars, 2 or 5 μm. m , Immunoprecipitation of GFP-myc ESB2 using GFP nanobody-coated beads followed by protein-blot analysis using an anti-HA antibody to detect the prey ( 6HA ESB3) and an anti-GFP antibody to detect the bait ( GFP-myc ESB2). IP, immunoprecipitation. The protein blots are representative of four independent experiments; a 6HA ESB3 single tagged cell line was used as a negative control. n , Violin plots depicting a comparative transcriptomic analysis between VEX2 ( n = 3), ESB2 ( n = 5), ESB3 ( n = 3) and ESAP1 ( n = 3) RNAi cell lines for ‘active’ ESAGs ; n values correspond to biological replicates. log 2 (FC), fold change in transcript abundance between RNAi (24 h post-induction) and the parental cell line. The violins span between minimum and maximum values, centre lines correspond to the mean and all datapoints are shown. Statistical significance was determined by one-way ANOVA followed by Tukey’s multiple-comparison test; **** P < 0.0001. Diagram in m created in BioRender; Faria, J. https://BioRender.com/oia6hdu (2026).

Article Snippet: The following primary antibodies were used: mouse α-myc (Millipore, clone 4A6, 1:10,000), mouse anti-Ty (BB2, Invitrogen, 1:5,000), mouse monoclonal anti-HA (Sigma, clone HA-7, 1:2,000), rabbit polyclonal anti-GFP (LifeTech, 1:2,000) and mouse α-EF1α (Millipore, clone CBP-KK1, 1:30,000).

Techniques: Fluorescence, Microscopy, Derivative Assay, Two Tailed Test, Knockdown, Staining, Immunoprecipitation, Negative Control, Comparison

Journal: Cells

Article Title: Host SNARE Proteins Mediate Lysosome and PVM Fusion to Support Plasmodium Liver Infection

doi: 10.3390/cells15070584

Figure Lengend Snippet: Localization of host SNARE proteins at the Plasmodium PVM during liver-stage infection. ( A – C ) Confocal live cell imaging of GFP-tagged SNARE proteins VAMP7-GFP ( A ), VAMP8-GFP ( B ), and Stx7-GFP ( C ) in P. berghei -infected HeLa cells expressing mCherry (parasite cytoplasm, red) at 6, 24, and 48 hpi. SNARE localization is shown in green. ( D – G ) Immunofluorescence analysis of Vti1B ( D ), VAMP7 ( E ), VAMP8 ( F ), and Stx7 ( G ) in HeLa cells fixed at 0.5, 1, and 1.5 hpi. SNAREs were detected with anti-GFP (green) or anti-Vti1B (green); the PVM was labeled with anti-UIS4 (red); and DAPI (blue) stained nuclei. ( H ) Localization of SNARE proteins (anti-Vti1B/anti-GFP in green) with the PVM marker UIS4 (red) at 24 hpi, visualized by expansion microscopy (5-fold expanded). Merged channels (yellow) highlight PVM-SNARE protein association. Nuclei were counterstained with DAPI (blue). Scale bars: 5 μm ( A – G ); 10 μm ( H ).

Article Snippet: The cells were then incubated in 10% FCS/PBS for 1 h, at room temperature with primary antibodies: Vti1B (mouse mAb 1:1000 (1:500 PS-ExM), BD Transduction Laboratories #611404, Allschwil, Switzerland), UIS4 (rabbit 1:1000, P. sinnis ; chicken 1:10000, Proteogenix, Schiltigheim, France), hLAMP1 (mouse mAb 1:1000 (1:500 PS-ExM), DSHB H4A3 (Iowa City, USA); rabbit pAb 1:1000, Cell Signaling #9091, Allschwill, Switzerland), GFP (rabbit pAb 1:1000, Origene TA100030, Herford, Germany; mouse mAb 1:1000, Roche AQ160, Basel, Switzerland), and α-tubulin (guinea pig pAb 1:500, ABCD AA345, Geneva, Switzerland).

Techniques: Infection, Live Cell Imaging, Expressing, Immunofluorescence, Labeling, Staining, Marker, Microscopy