Fig 2: Esrrg immunohistochemistry on sectioned embryonic mouse kidney. Photomicrographs of kidney sections counterstained with eosin (pink) and Esrrg is detected with NBT/BCIP (blue). 12.5 dpc staining in early-stage kidneys can only be seen in the peripheral uncondensed metanephric mesenchyme or stroma. 14.5, 15.5, 16.5 dpc: each kidney shows regionally restricted signal in specific ductal epithelia and in the peripheral mesenchyme. 18.5 dpc and P0 (first postnatal day): signal becomes restricted to the renal papilla and the developing ureter.

Fig 3: The LVST neuron group and the vicinal mediocaudal neuron group sharing the LVST group TF signature derive from different rhombomeric lineages. A, Expression of Maf/Esrrg/Phox2b confirmed in LVST neurons by direct combination with retrograde BDA labeling in the E13.5 mouse hindbrain. Maf/Esrrg/Phox2b triple-positive neurons were restricted to two clusters, the large LVST neuron group and the smaller, vicinal mediocaudal neuron group, seen in frontal (left) and lateral (right) views. Red circles, Maf+ Esrrg+ Phox2b+ BDA+ neurons (n = 306); blue triangles, Maf+ Esrrg+ Phox2b+ BDA- neurons (n = 59). Cells from every third section in this preparation counted. Dashed line indicates the midline. B–E, Transverse sections from an E14.5 b1r4-Cre x tdTomato mouse, triple-immunostained for Maf, Esrrg, and Phox2b, at the level of the vicinal mediocaudal neuron group (B, C) and the LVST neuron group (D, E). C, E, Separate confocal images of each fluorescent channel from within the dashed boxes shown in B and D, with red dots indicating cells colabeled for Maf/Esrrg/Phox2b. tdTomato staining is absent in the vicinal mediocaudal neuron group (C), indicating the origin of these neurons from outside of r4. The anti-Maf signal is not shown in B or D for clarity. B–E, Dorsal, up; lateral, left; midline, right.

Fig 4: ESRRG antagonizes Wnt signaling in GC. a Gene expression profile presented in a matrix format; each row represents an individual gene, and each column represents a transfected cell condition. In this matrix, red and green reflect relatively high and low expression, respectively, as indicated in the scale bar (log2-transformed scale). Genes associated with oncogenic potential and Wnt signaling-associated genes are listed. b qPCR analysis of Wnt signaling associated genes in GC cells (AGS and MKN28) after infection with Flag or Flag-ESRRG lentivirus. c, d Rescue experiments following the introduction of TCF4/LEF1. After infection of Flag or Flag-ESRRG, the indicated plasmids were transfected into AGS cells, and CCK8 and CFA assays were done. e, f The MKN45 cell line infected with Flag-ESRRG overexpression vector (ESRRG-OE) or Flag with TCF4/LEF1 was injected into female nude mice and the tumor volume was measured at the indicated time points (n = 5 per group). e Mice were sacrificed and tumor volumes and weights were measured. f IHC analysis from the mouse samples was then performed. g, h AGS cells were transiently transfected as indicated with the Top/Flash reporter (g) or CCND1 promoter (h) and the indicated constructs and reporter activity was measured using a luminometer. i Correlation of ESRRG and Wnt component gene expression (TCF7L2/TCF4, LEF1) in GSE29272 GC patient cohorts. Scatter plots of ESRRG and Wnt signaling genes in the GC cohorts are shown. Data represent the mean ± s.d. from three independent replicates. Student’s t-test was used to examine statistical significance (*p < 0.05, **p < 0.01, ***p < 0.005, ****p < 0.001)

Fig 5: Clinical relevance of ESRRG in GC patient prognosis. a Patients in the indicated GC cohorts were dichotomized by relatively high or relatively low ESSRG expression and were considered for plotting. b, c Establishment of gene expression profiles of ESRRG downstream targets. b Gene expression signature of ESRRG in control (Con.) or ESRRG lentivirus transduced AGS cells. Genes in the Venn diagram were selected by applying a two-sample Student’s t-test (p < 0.001) to compare Con. and ESRRG OE samples. b Overall scheme of prediction models and evaluation of predicted outcome based on a shared gene expression signature of ESRRG in GC cell lines. The ESRRG gene expression signature was used to form a series of classifiers that estimated the probability of how much the expression pattern of a particular patient with GC was similar to the shared signature; control (Con.) vs. ESRRG OE (over-expression). c Kaplan–Meier plots of OS of GC patients in the GSE26899 cohort merged with the GSE26901 and GSE14208 cohorts were used to predict outcome based on the Con. and ESRRG OE signatures as a classifier. The differences between groups were significant where indicated (log-rank test). SVM, support vector machines; LDA, linear discriminator analysis; 1NN, one nearest neighbors