Journal: Nature Communications

Article Title: Obesity-driven microbial GABA depletion promotes metabolic rewiring and colorectal cancer progression

doi: 10.1038/s41467-026-76079-1

Figure Lengend Snippet: A Venn diagram of proteins shared by DARTS and co-IP proteomics with GAP antibody, identifying PPP1CA as a GAP-binding protein. B Immunoblot of PPP1CA pulled down by GAP antibody (top) and reciprocal co-IP of GAP with PPP1CA antibody (bottom); The immunoblots shown were obtained from separate membranes. Representative of 2 independent experiments with similar results. C Molecular docking of GAP to PPP1CA: predicted binding interface (top) and interaction residues (bottom); GAP, green; PPP1CA, cyan. D Co-IP in 293 T cells expressing PPP1CA WT or mutant (R132A/R221A) and treated with GAP (IP, GAP antibody; immunoblot, PPP1CA); n = 2 independent experiments. E PPP1CA phosphatase activity in HT-29 treated with GAP; n = 3 independent experiments per group. F CCK-8 assay of HT-29 transfected with PPP1CA WT or mutant (R132A/R221A) and treated with GAP; n = 5 independent experiments per group. G Colony formation (same conditions); n = 5 independent experiments per group. H Western blot of phospho-YAP (P-YAP) and total YAP in HT-29 and RKO cells (PPP1CA WT or mutant; GAP or vehicle). I Immunofluorescence of YAP in intestinal tissues of Apc Min/+ mice after GABA. Blue arrows, cytoplasmic YAP; green arrows, nuclear YAP. Scale bar, 100 μm; zoomed-in images: 20 μm; representative of five mice per group with similar results. J Immunofluorescence of YAP in intestinal tissues of Apc Min/+ mice after B. ovatus colonization (arrows as in I). Scale bar, 100 μm; zoomed-in images: 20 μm; representative of five mice per group with similar results. K Immunofluorescence of YAP in tumor tissues from patients with obesity and CRC and patients with normal weight and CRC (arrows as in I ). Scale bar, 100 μm; zoomed-in images: 20 μm; representative of four patients per group with similar results. L RT-qPCR of Ctgf and Edn1 in colonic tissues of Apc Min/+ mice ( B. ovatus versus PBS); n = 3 mice per group. M RT-qPCR of Ctgf and Edn1 in colonic tissues of Apc Min/+ mice (GABA versus PBS); n = 3 mice per group. Statistics: two-sided unpaired Student’s t -test ( E , L , M ); two-sided one-way ANOVA with Tukey’s post hoc test ( F , G ). Source data are provided as a Source data file.

Article Snippet: The primary antibodies used included TPI1 (Proteintech Group, #10713-1-AP, 1:1000), GAPDH (Proteintech Group, #60004-1-Ig, 1:1000), Aldolase (Proteintech #11217-1-AP, 1:1000), β-Tubulin (Abcam, #ab6046, 1:2000), Phospho-YAP (Ser127) (Cell Signaling Technology, #13619), YAP (Abclonal, #A19134, 1:1000), PPP1CA (Abclonal, #A24288, 1:1000), and PGD (Proteintech Group, #22241-1-AP, 1:1000).

Techniques: Co-Immunoprecipitation Assay, Binding Assay, Western Blot, Expressing, Mutagenesis, Activity Assay, CCK-8 Assay, Transfection, Immunofluorescence, Quantitative RT-PCR