Figure 4.

myCAF activation induced by S100A8 is triggered via CD147-RhoA-ROCK-MLC2-MRTF-A pathway

(A) KEGG pathway enrichment analysis of distinct CAF subtypes.

(B) Spearman correlation between the S100A8 RNA level and the pathway signature score. The gray area represents 95% CI (n = 60).

(C) Western blot analysis of RhoA-associated protein markers in CAFs treated with indicated concentration of rS100A8 proteins.

(D) Western blot analysis of RhoA-associated and myCAF-related protein markers in CAFs transfected with siRNAs targeting RhoA, ROCK1, MLC2, or negative control and treated with rS100A8 proteins or PBS.

(E) Representative immunofluorescence images of MRTF-A nuclear translocation in CAFs treated with rS100A8 proteins or PBS. White arrows indicate MRTF-A nuclear translocation in cells. Scale bar, 100 μm.

(F) Quantification of the percentage of MRTF-A nuclear translocation in (E) (n = 6 biological replicates). Data are presented as mean ± SD. ∗∗∗∗p < 0.0001 of two-tailed Student’s t test.

(G) Western blot analysis of the indicated proteins in CAFs transfected with MRTF-A or control siRNA and treated with indicated concentration of rS100A8 proteins.

(H) Representative immunofluorescence images of CD147 and αSMA in control and CD147-knockout CAFs. Scale bar, 100 μm.

(I) Western blot analysis of the indicated proteins in control and CD147-knockout CAFs treated with indicated concentration of rS100A8 proteins.

(J) Western blot analysis of the indicated proteins in control and CD147-knockout CAFs cocultured with control or S100A8-overexpression KYSE450 cells.

Each assay for western blot had three biological repeats. See also Figures S4 and S5.