peroxiredoxin 3  (Bioss)

Journal: Nature Communications

Article Title: Trifunctional sphingomyelin derivatives enable nanoscale resolution of sphingomyelin turnover in physiological and infection processes via expansion microscopy

doi: 10.1038/s41467-024-51874-w

Figure Lengend Snippet: a , b TFSMs are enriched within the Golgi and mitochondria. HeLa cells were treated with TFSM 1 for 2 h ( a ) or 24 h ( b ). Samples were fixed and clicked to BODIPY-FL-DBCO (backbone) or Atto647N-azide (headgroup). Golgi and mitochondria were visualized by an anti-Golgi matrix protein 130 (GM130) and anti-Peroxiredoxin (Prx3) antibody, respectively. n = 1. Scale bars: 25 µm ( a ) or 5 µm ( b ). c , d FRET measurement of TFSM conversion. HeLa or human umbilical vein endothelial cells (HuVEC) were incubated with TFSM 1, TFSM 2, or BODIPY-FL-C 12 -SM for 2 h. Then, the compounds were removed, and cells were either treated with bSMase or left untreated for 3 h. Samples treated with TFSMs were fixed, clicked with BODIPY-FL-DBCO (backbone) and AlexaFluor TM 546 (AF)546-azide (headgroup) and FRET efficiency was determined by acceptor bleaching. Cells treated with BODIPY-FL-C 12 -SM were detached, and lipids were extracted by CHCl 3 :MeOH. The proportion of unmetabolized SM was determined by thin-layer chromatography. Scale bars: 25 µm. n = 3. Statistics: Two-way ANOVA and Šídák’s multiple comparisons ( d ). Bars represent means ± SD. n corresponds to biological replicates. Source data and detailed statistics are provided as a Source Data file.

Article Snippet: 7163670), 1:50 anti-Prx3 antibody (OriGene, Cat. No. TA322472) or 4 μg/ml anti-chlamydial HSP60 antibody (SantaCruz, Cat. No. sc-57840, Lot.

Techniques: Incubation, Thin Layer Chromatography