MUC4, a high–molecular weight transmembrane glycoprotein, is overexpressed in pancreatic cancer and is implicated in its pathogenesis. It is a heterodimeric protein containing a large extracellular, heavily glycosylated subunit, MUC4α, and a transmembrane growth factor–like subunit, MUC4β. In the present study, we have shown the interaction of human MUC4 with the receptor tyrosine kinase HER2 in pancreatic adenocarcinoma cells by reciprocal coimmunoprecipitation and cocapping studies. MUC4 colocalized with HER2 at the cell surface and in the cytoplasm. Silencing of MUC4 by transient or stable expression of MUC4-targeted short-interfering RNA led to the down-regulation of HER2 with a concomitant decrease in its phosphorylated form (pY1248-HER2). Further analyses revealed that the MUC4-knockdown–mediated decrease in HER2 expression occurred due to the drop in the stability of the receptor. In MUC4-knockdown pancreatic cancer cells, we also observed a reduced phosphorylation of the focal adhesion kinase and p42/44 mitogen-activated protein kinase, which are downstream effector proteins in HER2 signaling. Our findings add a new dimension to MUC4 function as a modulator of cell signaling and provide mechanistic evidence for its role in pancreatic cancer progression.