MUC4 is a large transmembrane type I glycoprotein that is overexpressed in pancreatic cancer (PC) and has been shown to be associated with its progression and metastasis. However, the exact cellular and molecular mechanism(s) through which MUC4 promotes metastasis of PC cells has been sparsely studied. Here we showed that the NIDO domain of MUC4, which is similar to the G1-domain present in the nidogen or entactin (an extracellular matrix protein), contributes to the protein-protein interaction property of MUC4. By this interaction, MUC4 promotes breaching of basement membrane integrity, and spreading of cancer cells. These observations are corroborated with the data from our study using an engineered MUC4 protein without the NIDO domain, which was ectopically expressed in the MiaPaCa PC cells, lacking endogenous MUC4 and nidogen protein. The in vitro studies demonstrated an enhanced invasiveness of MiaPaCa cells expressing MUC4 (MiaPaCa-MUC4) compared to vector-transfected cells (MiaPaCa-Vec; p=0.003) or cells expressing MUC4 without the NIDO domain (MiaPaCa-MUC4-NIDO Δ; p=0.03). However, the absence of NIDO-domain has no significant role on cell growth and motility (p=0.93). In the in-vivo studies, all the mice orthotopically implanted with MiPaCa-MUC4 cells developed metastasis to the liver as compared to MiaPaCa-Vec or the MiaPaCa-MUC4-NIDO Δ group, hence, supporting our in vitro observations. Additionally, a reduced binding (p=0.0004) of MiaPaCa-MUC4-NIDO Δ cells to the fibulin-2 coated plates compared to MiaPaCa-MUC4 cells indicated a possible interaction between the MUC4-NIDO domain and fibulin-2, a nidogen-interacting protein. Furthermore, in PC tissue samples, MUC4 colocalized with the fibulin-2 present in the basement membrane. Altogether, our findings demonstrate that the MUC4-NIDO domain significantly contributes to the MUC4-mediated metastasis of PC cells. This may be partly due to the interaction between the MUC4-NIDO domain and fibulin-2.