Journal: Cell

Article Title: Structural Basis of Membrane Protein Chaperoning through the Mitochondrial Intermembrane Space

doi: 10.1016/j.cell.2018.10.039

Figure Lengend Snippet:

Article Snippet: 35 S-methionine , Perkin Elmer , Cat. #NEG009T005MC.

Techniques: Produced, Virus, Recombinant, Labeling, Dispersion, Software

Journal: Communications Biology

Article Title: Gasdermin-D pores induce an inactivating caspase-4 cleavage that limits IL-18 production in the intestinal epithelium

doi: 10.1038/s42003-025-08183-9

Figure Lengend Snippet: Human duodenal organoids were trypsinised into single cells and seeded onto culture plates pre-coated with 1:40 Matrigel:PBS. Cells were infected with S. flexneri ∆ ospc3 at an MOI of 100 for 4 h. CFUs in infected cells were enumerated ( a ), PI uptake was monitored using incucyte live cell imager and PI uptake was normalised per area confluency ( b ), images of monolayers taken 4 h post infection taken at 20x (i) scramble, uninfected; (ii) scramble, Shigella flexneri ∆ ospc3, (iii) GSDMD KD uninfected, (iv) GSDMD KD Shigella flexneri ∆ ospc3. Scale bars (indicting 200 micrometres) are depicted for each micrograph ( c ). Caspase-4 was measured in western blot of whole cell lysates ( d ), IL-18 secretion into the supernatant was measured by ELISA on cell free supernatants ( e ). A schematic of caspase-4 regulation ( f ). In ( f ) (i) LPS-induced caspase-4 activation (1) leads to activation of GSDMD and IL-18 (2). GSDMD pores lead to caspase-4 cleavage (3) and this stops IL-18 processing (4). In ( f )(ii), during conditions of reduced GSDMD –mediated plasma pore formation, caspase-4 activation (1) leads to IL-18 processing. A lack of efficient pore formation reduces inhibitory feedback on caspase-4 (3). This leads to continued activity of caspase-4 on IL-18 and IL-18 hypersecretion (4). All experiments are representative of at least 3 experiments. Data displayed as mean ± S.E.M. * p < 0.05, **** p < 0.001. Panel ( f ) created with Biorender.

Article Snippet: Tissue was minced until able to be passed through a 1 mL pipette tip then incubated in 2 mg/mL collagenase at 37 o C for 40 min. Crypt units were dissociated by vigorous pipetting then washed 3x in PBS containing 10% FBS and plated into 40 μL Matrigel domes (Cultrex PathClear reduced growth factor BME, type 2; R&D Systems) in a 24 well plate.

Techniques: Infection, Western Blot, Enzyme-linked Immunosorbent Assay, Activation Assay, Clinical Proteomics, Activity Assay