Fig. 1: MST3 inhibits growth and metastasis of CRC.

a Representative immunohistochemical images of MST3 in human colorectal peri-tumor and tumor tissues from CRC patients. Scores of MST3 expression levels were quantified. n = 59. Scale bar: 50 µm. P = 3.04E-11. b qRT-PCR analysis for MST3 in human peri-tumor and tumor tissues from CRC patients. n = 10. P = 0.03. c Representative immunohistochemical images of Mst3 in mouse colon peri-tumor and tumor tissues from AOM-DSS mouse model. n = 3. Scale bar: 50 µm. d Gross images of AOM-DSS mouse colon tumors from control (Ctrl) and Mst3^cKO mice. Tumor area (P = 1.58E-12) and the number of tumors (P = 0.002) per mouse were quantified. Ctrl: n = 50 tumors from 6 mice. Mst3^cKO: n = 73 tumors from 5 mice. e Representative histological images of AOM-DSS colon tumors from Ctrl and Mst3^cKO mice. n = 3. Scale bar: 2 mm. f Anchorage-independent growth of HCT116 cells transfected with NC or siMST3. Colony formation rate was determined. Colonies grew for 5 weeks. NC, n = 27 fields from 3 independent samples; siMST3, n = 38 fields from 3 independent samples. Scale bar: 10 μm. P = 1.94E-08. g Schematic diagram of constructs of human MST3 kinase-dead mutant plasmid (Lysine at position 53 was mutated to Alanine) and kinase persistent phosphorylation mutant plasmid (Threonine at position 178 was mutated to Glutamate). h HA-labeled pcDNA3.1 empty vector, pcDNA3.1-MST3^WT, pcDNA3.1-MST3^K53A and pcDNA3.1-MST3^T178E were transfected into APKS mouse tumor organoids, respectively. Organoid areas were quantified at the indicated time points. 4 h, n = 50 fields from 3 independent samples; 52 h, n = 53 fields from 3 independent samples. Scale bar: 250 μm. i Schematic diagram of the cecal orthotopic injection mouse model using luciferase-labelled HCT116 cells. j HCT116 cells with stable knockdown of MST3 (shMST3) transfected with pcDNA3.1, pcDNA3.1-MST3^WT, pcDNA3.1-MST3^K53A or pcDNA3.1-MST3^T178E plasmids were injected into the cecum of NOG mice, and bright field images of primary tumor growth and quantification of tumor weight were recorded. n = 3 mice per group. P = 0.032; P = 0.027; P = 0.009. k Representative IVIS luciferase images of CRC liver metastases in NOG mice injected with HCT116 cells into the cecum. Data are presented as the mean ± SD. *P < 0.05; **P < 0.01; ***P < 0.001. Statistical analysis in panel h was performed with one-way ANOVA followed by Tukey’s test, and the rest were performed with two-tailed unpaired Student’s t-test.