Fig 2: Integrin ß1 binds to NF-?B p105 and promotes the NF-?B p50/65 pathway. a The protein?protein interaction network of the Sema7a, integrin ß1 and NF-?B protein families was predicted by the Retrieval of Interacting Genes Database. b Coimmunoprecipitation analysis of protein interactions among integrin ß1 and NF-?B p105 in liver tissues from 8-week-old male wild-type and Sema7a.R145W homozygous mice. The higher band is NF-?B p105 (105 kDa) and the lower band is NF-?B p55 (55 kDa). c Schematic showing the primary structure and truncation of NF-?B p105. d and e Coimmunoprecipitation analysis of His-tagged NF-?B p105 and Flag-tagged integrin ß1 in HEK293 cells. f, g Co-immunoprecipitation analysis of His-tagged NF-?B p55 and Flag-tagged integrin ß1 in HEK293 cells. h Coimmunoprecipitation analysis of the protein interaction between Sema7a and NF-?B p105 in primary mouse hepatocytes after integrin ß1 silencing (n = 3). i, j Western blotting analysis of the relative levels of integrin ß1, NF-?B p105, p- NF-?B p50/NF-?B p50, p- NF-?B p65/NF-?B p65, p-I?B/I?B, TNFa and IL1ß protein expression in primary hepatocytes following integrin ß1 silencing (n = 3). Phosphorylation levels were measured by the phosphor/total protein ratio. The difference among the groups was determined by one-way ANOVA with Tukey’s post hoc tests or by Kruskal?Wallis test with Dunn’s post hoc test analysis. *Means p < 0.05

Fig 3: SEMA7A is elevated in hepatocellular carcinoma. a The mRNA level of SEMA7A in liver hepatocellular carcinoma from TCGA. b and c The mRNA levels of SEMA7A and integrin ß1 in four tumour grades. d Characteristic images of liver tissue from wild-type and hepatocellular carcinoma mice. e Representative images of H&E staining in wild-type and hepatocellular carcinoma. f, g Western blotting analysis of the relative levels of Sema7a, integrin ß1, NF-?B p105, p- NF-?B p50/NF-?B p50, p- NF-?B p65/NF-?B p65, p-I?B/I?B, TNFa and IL1ß protein expression in the livers of wild-type (n = 5) and hepatocellular carcinoma mice (n = 5). h The relative levels of mRNA transcripts of the genes for SEMA7A in wild-type (n = 5) and hepatocellular carcinoma mice (n = 5). i Coimmunoprecipitation analysis of protein interactions among integrin ß1 and NF-?B p105 in liver tissues from wild-type and hepatocellular carcinoma mice. The data were analysed by the independent-samples Student’s t test. *Means p < 0.05 versus wild-type mice

Fig 4: Ectopic expression of SEMA7A promotes NF-?B p105 procession and activates the NF-?B p50/p65 pathway. a and b Relative Sema7a expression in HCC cell lines (HepG2, Huh7, PLC5, and Hep3B) compared with an immortalized normal human hepatic cell line (L02). c Sema7a overexpression efficiency and relative NF-?B1, TNFa and IL1ß mRNA levels in transfected HepG2 cell lines (n = 3). d and e Western blotting analysis of the relative levels of Sema7a, integrin ß1, NF-?B p105, NF-?B p50, p- NF-?B p65/NF-?B p65, p-I?B/I?B, TNFa and IL1ß protein expression in transfected HepG2 cell lines following integrin ß1 silencing (n = 3). Phosphorylation levels were measured by the phosphor/total protein ratio. The qPCR data were analysed by the independent-samples Student’s t test and differences among the groups were determined by one-way ANOVA with Tukey’s post hoc tests or by Kruskal?Wallis test with Dunn’s post hoc test analysis. *Means p < 0.05

Fig 5: The NF-?B pathway is activated in Sema7aR145W mouse liver and primary mouse hepatocytes. a–c Western blotting analysis of the relative protein levels of NF-?B p105, p- NF-?B p50/NF-?B p50, p- NF-?B p65/NF-?B p65 and p-I?B/I?B in the livers of wild-type (n = 4) and Sema7aR145W homozygous male mice (n = 5) and b, d in primary hepatocytes from wild-type (n = 3) and Sema7aR145W homozygous (n = 3) male mice. Phosphorylation levels were measured by the phosphor/total protein ratio. e Representative IF staining showing p-NF-?B p65 Ser529 (red) and DAPI (blue). Scale bars: 50 µm in liver sections from wild-type and Sema7aR145W homozygous male mice. f The relative levels of mRNA transcripts of the genes for NF-?B p105 in wild-type (n = 4) and Sema7aR145W homozygous male mice (n = 5) and g in primary hepatocytes from wild-type (n = 3) and Sema7aR145W homozygous (n = 3) male mice. The data were analysed by the independent-samples Student’s t test. * means p < 0.05 versus wild-type mice