Fig 2: Methodological verification of HSPE1 in individual urine specimens from BC and hernia patients by antibody-based analysis(A) Expression of HSPE1 in individual clinical urine specimens from BC and hernia patients, detected by Western blot analysis using an anti-HSPE1 antibody. A pooled urine sample prepared from BC and hernia patients was used as control for comparison of multiple gels. Data were quantified using Image J software. (B–D) Results of statistical analyses of HSPE1 expression in urine from hernia and BC patients by ELISA. (B) Differences in HSPE1 concentrations in urine specimens from hernia (n = 142), BC (n = 59), hematuria (n = 67) and UTI (n = 59) patients. (C) Difference in HSPE1 concentration in urine specimens from hernia (n = 142), low-grade BC (n = 42) and high-grade BC (n = 84) patients. (D) Difference in HSPE1 concentration in urine specimens from hernia (n = 142), early-stage BC (n = 98), and advanced-stage BC (n = 28) patients. Differences between groups were determined using the Mann-Whitney test.

Fig 3: NPM1, HSPE1, and HSPB1 were both O-GlcNAc-modified and phospho-modified. a ETD mass spectra of peptides derived from NPM1, HSPE1, and HSPB1. b The O-GlcNAcylation and phosphorylation levels of endogenous NPM1, HSPE1, and HSPB1 were measured in HB tissues and normal tissues by immunoprecipitation and western blot analysis. All experiments were repeated three times

Fig 4: Fhit interacts with Fdxr, HSP10, and HSP60.a GST pull-down experiment using a protein lysate from H1299D1 cells transfected with HA-FDXR which was incubated with GST and GST-Fhit protein bound to glutathione agarose resin, and protein complexes eluted and separated on acrylamide gel for detection with antisera against HA, HSP60, and Fhit. b Lysates were prepared from DSP-cross-linked H1299D1 cells (Pon A induced for Fhit expression) or from HCT116 cells (expressing endogenous Fhit) and used in IP experiments with the indicated antisera. Complexes were separated on acrylamide gel and proteins were detected with HSP60, Fhit, or HSP10 antisera. c The association of Fhit and Fdxr in HCT116 cells using the Duolink in situ proximity ligation assay (PLA). PLA signals are shown in red and nuclei in blue. PLA signals for Fhit/Fdxr are mostly confined in the cytoplasm. Negative controls with the Fdxr or Fhit antibody alone show no PLA signal. Scale bar represents 10 µm