p aeruginosa cmcc 10104  (ATCC)

Journal: bioRxiv

Article Title: GPLD1 functions as a scavenger chaperone mediating lysosomal degradation of extracellular aberrant proteins

doi: 10.1101/2025.07.16.665076

Figure Lengend Snippet: ( A ) Catalytic activity of GPLD1 is not required for its internalization. Wild-type (WT) GPLD1-RFP-GFP or catalytic mutants (H133N, H158N) were preincubated with luciferase (42°C for 30 min) or ALFA-SAA1 (37°C for 3 h) in serum-free medium. HEK293 cells were treated with medium containing GPLD1-RFP-GFP with or without substrate for 18 h and analyzed by flow cytometry. The bar graph shows relative fluorescence intensity per cell normalized to untreated controls (n = 3). Data are presented as mean ± SEM. ( B ) Confirmation of GPAA1 knockout (KO) by immunoblotting wild-type and GPAA1 KO HeLa cell lysates with anti-GPAA1 antibody. ( C ) GPAA1 KO cells fail to mature GPI-anchored proteins. Wild-type and GPAA1 KO HeLa cells expressing GFP-GPI were analyzed by confocal microscopy. Scale bar, 10 µm. ( D ) GPLD1 internalization is not impaired in GPAA1 KO cells. Wild-type and GPAA1 KO HeLa cells were treated as in (A) and analyzed by flow cytometry.

Article Snippet: Rabbit polyclonal anti-β-actin (cat# 20536-1-AP) and anti-GPAA1 (cat# 10104-1-AP) antibodies were obtained from Proteintech.

Techniques: Activity Assay, Luciferase, Flow Cytometry, Fluorescence, Knock-Out, Western Blot, Expressing, Confocal Microscopy