Journal: The Cell Surface

Article Title: Pap1 is an adhesin involved in the interaction of Sporothrix schenckii and Sporothrix brasiliensis with the host

doi: 10.1016/j.tcsw.2025.100164

Figure Lengend Snippet: In silico docking between Pap1 and fibronectin. In the upper panel, Sporothrix schenckii Pap1 is in green, and fibronectin is in orange. In the lower panel, Sporothrix brasiliensis Pap1 is in orange and fibronectin is in blue. In both panels, the surface pocket involved in binding is in grey, while the amino acids interacting with the ligand are in red. Both images were generated with PyMol. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: Panels A and C are results generated with S. schenckii yeast-like cells (WT, 1099–18 ATCC MYA 4821 strain); while panels B and D are results obtained with S. brasiliensis strains (WT, 5110 ATCC MYA 4823 strain).

Techniques: In Silico, Binding Assay, Generated

Journal: The Cell Surface

Article Title: Pap1 is an adhesin involved in the interaction of Sporothrix schenckii and Sporothrix brasiliensis with the host

doi: 10.1016/j.tcsw.2025.100164

Figure Lengend Snippet: Adhesion to extracellular matrix components and HeLa cells of Sporothrix schenckii and Sporothrix brasiliensis PAP1 -silenced strains. The indicated extracellular matrix component was used to coat 96-well plates, then yeast-like cells were added, unbound cells were removed by extensive washing, and adherent cells were detected by ELISA with a primary anti-rHsp60 antibody. Control refers to wells coated with bovine serum albumin. Panel A contains data generated with S. schenckii, and WT is the 1099–18 ATCC MYA 4821 strain. Panel B shows results with S. brasiliensis cells, and the WT is the 5110 ATCC MYA 4823 strain. Alternatively, for both panels, 1 × 10 6 HeLa cells were placed per well, incubated 24 h at 37 °C and 5 % ( v /v) CO 2 , and used in the adhesion assays. Results are means ± SD of three biological replicates performed in duplicate. The Dunnett's test and then the unpaired t-test were used for data analysis. In both panels, * P < 0.05 when compared to WT or strains HSB1 and HSB2.

Article Snippet: Panels A and C are results generated with S. schenckii yeast-like cells (WT, 1099–18 ATCC MYA 4821 strain); while panels B and D are results obtained with S. brasiliensis strains (WT, 5110 ATCC MYA 4823 strain).

Techniques: Enzyme-linked Immunosorbent Assay, Control, Generated, Incubation

Journal: The Cell Surface

Article Title: Pap1 is an adhesin involved in the interaction of Sporothrix schenckii and Sporothrix brasiliensis with the host

doi: 10.1016/j.tcsw.2025.100164

Figure Lengend Snippet: Biofilm formation in Sporothrix schenckii and Sporothrix brasiliensis PAP1 -silenced strains. Biofilms were generated in microplates, using yeast-like cells. Non-adherent cells were removed, and biofilms were allowed to mature for 24 h at 37 °C. Biomass was stained with crystal violet and quantified by spectrophotometry. Panel A shows data with S. schenckii cells, and the WT is 1099–18 ATCC MYA 4821 strain. Panel B contains results obtained with S. brasiliensis cells, and the WT is the 5110 ATCC MYA 4823 strain. Data are means ± SD of three biological replicates. Results were analyzed with Dunnett's test and then the t-test. * P < 0.05 when compared to WT.

Article Snippet: Panels A and C are results generated with S. schenckii yeast-like cells (WT, 1099–18 ATCC MYA 4821 strain); while panels B and D are results obtained with S. brasiliensis strains (WT, 5110 ATCC MYA 4823 strain).

Techniques: Generated, Staining, Spectrophotometry

Journal: The Cell Surface

Article Title: Pap1 is an adhesin involved in the interaction of Sporothrix schenckii and Sporothrix brasiliensis with the host

doi: 10.1016/j.tcsw.2025.100164

Figure Lengend Snippet: Cell wall rhamnose and mannose content in Sporothrix schenckii and Sporothrix brasiliensis wild-type, control, and PAP1 -silenced mutant strains. Yeast-like cells were homogenized, cell walls were isolated, cleansed, acid-hydrolyzed, and the resulting monosaccharides were separated and quantified by high-performance anion-exchange chromatography with pulsed amperometric detection. In A, results generated with S. schenckii cells. WT, 1099–18 ATCC MYA 4821 strain. In B, results obtained with S. brasiliensis strains. WT, 5110 ATCC MYA 4823 strain. For both panels, data are means ± SD of three biological replicates. Results were analyzed with Dunnett's test and then the t -test. * P < 0.05 when compared to WT, or control strains (HSS67 and HSS68 in panel A; HSB28 and HSB29 in panel B).

Article Snippet: Panels A and C are results generated with S. schenckii yeast-like cells (WT, 1099–18 ATCC MYA 4821 strain); while panels B and D are results obtained with S. brasiliensis strains (WT, 5110 ATCC MYA 4823 strain).

Techniques: Control, Mutagenesis, Isolation, Chromatography, Generated

Journal: The Cell Surface

Article Title: Pap1 is an adhesin involved in the interaction of Sporothrix schenckii and Sporothrix brasiliensis with the host

doi: 10.1016/j.tcsw.2025.100164

Figure Lengend Snippet: Cytokine stimulation by human peripheral blood mononuclear cells. Yeast-like cells and human peripheral blood mononuclear cells were coincubated for 24 h, and the secreted cytokines were quantified by ELISA. Panels A , C , and E are results generated with S. schenckii yeast-like cells (WT, 1099–18 ATCC MYA 4821 strain), while panels B , D , and F are results obtained with S. brasiliensis strains (WT, 5110 ATCC MYA 4823 strain). None, human cells preincubated with 5 μg mL −1 polymyxin B; Anti-MR, human cells preincubated with 5 μg mL −1 polymyxin B and 10 μg mL −1 anti-mannose receptor antibody. Anti-TLR4, human cells preincubated with 5 μg mL −1 polymyxin B and 10 μg mL −1 anti-TLR4. Data are means ± SD obtained with samples from eight donors, each assayed in duplicate wells. Results were analyzed with Dunnett's test and then the Mann-Whitney U test. * P < 0.05 when compared to strains WT, HSS67, or HSS68 (panels A , C , and E ). * P < 0.05 when compared to strains WT, HSB28, or HSB29 (panels B , D , and F ). † P < 0.05 when compared to the “None” group from the same strain.

Article Snippet: Panels A and C are results generated with S. schenckii yeast-like cells (WT, 1099–18 ATCC MYA 4821 strain); while panels B and D are results obtained with S. brasiliensis strains (WT, 5110 ATCC MYA 4823 strain).

Techniques: Enzyme-linked Immunosorbent Assay, Generated, MANN-WHITNEY

Journal: The Cell Surface

Article Title: Pap1 is an adhesin involved in the interaction of Sporothrix schenckii and Sporothrix brasiliensis with the host

doi: 10.1016/j.tcsw.2025.100164

Figure Lengend Snippet: Phagocytosis of Sporothrix schenckii and Sporothrix brasiliensis PAP1 -silenced strains. Yeast-like cells were labeled with Acridine Orange and used to interact with human monocyte-derived macrophages for 2 h at 37 °C and 5 % (v/v) CO 2 . Then, macrophages were collected and analyzed by flow cytometry. Macrophages that were interacting with at least one red fluorescent yeast-like cell were included in the analysis. None, human cells preincubated with 5 μg mL-1 polymyxin B. Anti-MR, human cells preincubated with 5 μg mL −1 polymyxin B and 10 μg mL −1 anti-mannose receptor antibody. Anti-TLR4, human cells preincubated with 5 μg mL −1 polymyxin B and 10 μg mL −1 anti-TLR4. Panel A , results generated with S. schenckii yeast-like cells (WT, 1099–18 ATCC MYA 4821 strain); while in panel B are results obtained with S. brasiliensis strains (WT, 5110 ATCC MYA 4823 strain). Data are means ± SD obtained with samples from eight donors, each assayed in duplicate wells. Results were analyzed with Dunnett's test and then the Mann-Whitney U test. * P < 0.05 when compared to strains WT, HSS67, or HSS68 (panel A ). * P < 0.05 when compared to strains WT, HSB28, or HSB29 (panel B ). † P < 0.05 when compared to the “None” group from the same strain. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: Panels A and C are results generated with S. schenckii yeast-like cells (WT, 1099–18 ATCC MYA 4821 strain); while panels B and D are results obtained with S. brasiliensis strains (WT, 5110 ATCC MYA 4823 strain).

Techniques: Labeling, Derivative Assay, Flow Cytometry, Generated, MANN-WHITNEY

Journal: The Cell Surface

Article Title: Pap1 is an adhesin involved in the interaction of Sporothrix schenckii and Sporothrix brasiliensis with the host

doi: 10.1016/j.tcsw.2025.100164

Figure Lengend Snippet: Cytokine stimulation by human monocyte-derived macrophages. Yeast-like cells and monocyte-derived macrophages were coincubated for 24 h, and the secreted cytokines were quantified by ELISA. Panels A and C are results generated with S. schenckii yeast-like cells (WT, 1099–18 ATCC MYA 4821 strain); while panels B and D are results obtained with S. brasiliensis strains (WT, 5110 ATCC MYA 4823 strain). None, human cells preincubated with 5 μg mL −1 polymyxin B. Anti-MR, human cells preincubated with 5 μg mL −1 polymyxin B and 10 μg mL −1 anti-mannose receptor antibody. Anti-TLR4, human cells preincubated with 5 μg mL −1 polymyxin B and 10 μg mL −1 anti-TLR4. Data are means ± SD obtained with samples from eight donors, each assayed in duplicate wells. Results were analyzed with Dunnett's test and then the Mann-Whitney U test. * P < 0.05 when compared to strains WT, HSS67, or HSS68 (panels A and C ). * P < 0.05 when compared to strains WT, HSB28, or HSB29 (panels B and D ). cells. † P < 0.05 when compared to the “None” group from the same strain.

Article Snippet: Panels A and C are results generated with S. schenckii yeast-like cells (WT, 1099–18 ATCC MYA 4821 strain); while panels B and D are results obtained with S. brasiliensis strains (WT, 5110 ATCC MYA 4823 strain).

Techniques: Derivative Assay, Enzyme-linked Immunosorbent Assay, Generated, MANN-WHITNEY

Journal: The Cell Surface

Article Title: Pap1 is an adhesin involved in the interaction of Sporothrix schenckii and Sporothrix brasiliensis with the host

doi: 10.1016/j.tcsw.2025.100164

Figure Lengend Snippet: Killing curves of Galleria mellonella larvae inoculated with Sporothrix schenckii and Sporothrix brasiliensis WT, control, and PAP1 -silenced strains. The G. mellonella larvae were inoculated with 10 μL of a yeast-like cell suspension at 1 × 10 7 cells mL -1, and survival was monitored daily for two weeks. Mortality is shown in Kaplan–Meier plots. In A, data generated with S. scehcnkii strains; while in B are shown the results with S. brasiliensis strains. Data were analyzed with the Log-rank test. In both panels, curves generated with the five PAP1 -silenced strains were not significantly different among themselves but were significantly different from the curves generated with the WT or control strains ( P < 0.05). In A, WT, 1099–18 ATCC MYA 4821 strain. Control strains are HSS67 and HSS68. In B, WT, 5110 ATCC MYA 4823 strain. Control strains are HSB28 and HSB29. For each strain, a total of 30 larvae were included in the study. In both panels, PBS, a larval group inoculated only with phosphate saline buffer.

Article Snippet: Panels A and C are results generated with S. schenckii yeast-like cells (WT, 1099–18 ATCC MYA 4821 strain); while panels B and D are results obtained with S. brasiliensis strains (WT, 5110 ATCC MYA 4823 strain).

Techniques: Control, Suspension, Generated, Saline

Journal: Molecular Biology of the Cell

Article Title: Polarized anionic phospholipids and exocytosis are implicated in the polarized recruitment of budding yeast AP180, an endocytic initiator

doi: 10.1091/mbc.E24-10-0446

Figure Lengend Snippet: Snc2 interacts with the endocytic machinery in a complex manner. Three-minute movies acquired using a Nikon confocal microscope equipped with an NSPARC detector (see Materials and Methods) of live cells expressing GFP-Snc2 (green) and Sla1-mScarlet (magenta), both from the endogenous promoter and locus from the endogenous promoter and locus in an snc1∆ background. Moving arrowheads track distinct Snc2-positive compartments.

Article Snippet: The yeast synaptobrevins Snc1/2 thus provide a unique opportunity to study the earliest stages of CME and better characterize a yeast CME cargo, a task that has remained difficult and yielded mixed results in past studies ( Toshima et al. , 2006 ; Carroll et al. , 2009 ).

Techniques:

Journal: Molecular Biology of the Cell

Article Title: Polarized anionic phospholipids and exocytosis are implicated in the polarized recruitment of budding yeast AP180, an endocytic initiator

doi: 10.1091/mbc.E24-10-0446

Figure Lengend Snippet: Snc2 interacts with the endocytic machinery in a complex manner. Three-minute movies acquired using a Nikon confocal microscope equipped with an NSPARC detector (see Materials and Methods) of live cells expressing GFP-Snc2 (green) and Sla1-mScarlet (magenta), both from the endogenous promoter and locus from the endogenous promoter and locus in an snc1∆ background. Moving arrowheads track distinct Snc2-positive compartments.

Article Snippet: The yeast synaptobrevins Snc1/2 thus provide a unique opportunity to study the earliest stages of CME and better characterize a yeast CME cargo, a task that has remained difficult and yielded mixed results in past studies ( Toshima et al. , 2006 ; Carroll et al. , 2009 ).

Techniques:

Journal: Molecular Biology of the Cell

Article Title: Polarized anionic phospholipids and exocytosis are implicated in the polarized recruitment of budding yeast AP180, an endocytic initiator

doi: 10.1091/mbc.E24-10-0446

Figure Lengend Snippet: Yap1801/2 is recruited to the daughter cortex by Snc1/2 and negatively charged lipids. A. Sequence alignment for human CALM and S. cerevisiae Yap1802. Red fill indicates direct matches, and red characters indicate shared residue characteristics . B. Alphafold2 multimer analysis of the ANTH domain of Yap1802 and Snc2, with key residues highlighted in red. C. Maximum intensity projections from 6-min widefield microscopy movies of cells expressing various Yap1802 wild-type or point-mutant alleles, 3x PM refers to the K21E K23E L203S triple point mutant. D. Circumferential kymographs made from 6-min widefield microscopy movies of Yap1802-GFP point mutants in medium-budded daughter cells. Temporal resolution is 2 s. E. Mean intensities of Yap1802-GFP point mutant line profiles from daughter cell cortexes as in C. Right to left, n = 35, 32, 23, 22 F. Quantification of Yap1802-GFP point mutant lifetimes. Right to left, n = 127, 129, 106, 97. G. Maximum intensity projections from 3-min Airyscan confocal microscopy time-lapse movies of cells expressing wild-type Yap1802 or point mutants and mScarlet-Snc2. Temporal resolution of movies was ∼3.7 s. H. Mean intensities of mScarlet-Snc2 line profiles from mother cell cortexes as in G. Right to left, n = 17, 15, 17, 14. Groups in E, F, and H were compared using one-way ANOVA. Center and error bars are mean, and 95% confidence interval, *** indicates 0.001 > p > 0.0001, and **** indicates p < 0.0001.

Article Snippet: The yeast synaptobrevins Snc1/2 thus provide a unique opportunity to study the earliest stages of CME and better characterize a yeast CME cargo, a task that has remained difficult and yielded mixed results in past studies ( Toshima et al. , 2006 ; Carroll et al. , 2009 ).

Techniques: Sequencing, Residue, Microscopy, Expressing, Mutagenesis, Confocal Microscopy

Journal: Molecular Biology of the Cell

Article Title: Polarized anionic phospholipids and exocytosis are implicated in the polarized recruitment of budding yeast AP180, an endocytic initiator

doi: 10.1091/mbc.E24-10-0446

Figure Lengend Snippet: A. Model for how interaction between synaptobrevin-like v-SNARES Snc1/2 and the CME adaptor proteins Yap1801/2 contributes to CME site initiation and links endocytosis to polarized exocytosis. Cdc42-nucleated actin cables enable directional transport of vesicles toward the daughter bud tip, with fusion near the bud tip assisted by the exocyst vesicle tethering complex. This localized secretion enriches the daughter plasma membrane in Snc1/2, which, in concert with anionic phospholipids, recruits Yap1801/2 to internalize Snc1/2 via CME. B. Model for Snc1/2 and anionic phospholipid coincidence detection, enabling robust Yap1801/2 recruitment. Each Yap1802 mutant displays defects in recruitment when compared with wild-type, an effect most pronounced in the triple point mutant. Red dots represent the rough location of each corresponding point mutation on the Yap1802 ANTH domain protein structure. Created with BioRender.com .

Article Snippet: The yeast synaptobrevins Snc1/2 thus provide a unique opportunity to study the earliest stages of CME and better characterize a yeast CME cargo, a task that has remained difficult and yielded mixed results in past studies ( Toshima et al. , 2006 ; Carroll et al. , 2009 ).

Techniques: Clinical Proteomics, Membrane, Mutagenesis