Fig. 4

MUC5AC physically interacts with CD44 to accelerate cellular signaling via Src. a Schematic representation of conditioned media (CM) collected from parental and KO clones (Clone-1 and -2) of HCT-8 and LS174T cell lines. b MUC5AC concentration is measured before or after CM preparation by ELISA. c and d Immunoblotting of CD44 from KO clones (Clone-1 and -2) treated with CM collected from parental cell lines (HCT-8 and LS174T) showed higher expression of CD44 in the presence of CM compared with untreated cells. No difference in CD44 expression was observed in KO clones supplemented with conditioned medium from KO clones. e Significantly higher expression of CD44 was observed in tumor patients (N = 275) compared with healthy controls (N = 41) in TCGA-COAD (^*p < 0.05). f MUC5AC and CD44 correlated positively in the TCGA-COAD dataset. g Immunoblot showed a physical interaction of MUC5AC-CD44 in CRC cell lines. h Both HCT-8 and LS174T cell lines co-stained with MUC5AC and CD44 antibodies. FITC-labelled and Alexa Fluor-labelled secondary antibodies were used for MUC5AC (Green) and CD44 (Red) staining, respectively. DAPI was used for nuclei staining. i Interaction of MUC5AC with CD44 mediates activation of various down-stream signaling molecules assessed in whole cell lysates isolated from parental and KO clones (Clone-1 and -2) of HCT-8. j Transient knock-down of CD44 altered its down-stream signaling molecules