We and others previously showed that protein kinase D1 (PKD1) is downregulated in several cancers including prostate; interacts with E-cadherin, a major cell adhesion epithelial protein; and causes increased cell aggregation and decreased motility of prostate cancer cells. In this study, we show that PKD1 complexes with β3-integrin, resulting in activation of mitogen-activated protein kinase/extracellular signal-regulated kinase (ERK) kinase–ERK pathway, which causes increased production of matrix metalloproteinase (MMP)-2 and MMP-9, that is associated with shedding of soluble 80 kDa E-cadherin extracellular domain. Interestingly, decreased cell proliferation following PKD1 transfection was rescued by MMP-2 and MMP-9 inhibitors and augmented by recombinant MMP-2 (rMMP-2) and rMMP-9 proteins, suggesting an antiproliferative role for MMPs in prostate cancer. Translational studies by in silico analysis of publicly available DNA microarray data sets show a significant direct correlation between PKD1 and MMP-2 expression in human prostate tissues. The study shows a novel mechanism for antiproliferative effects of PKD1, a protein of emerging translational interest in several human cancers, through increased production of MMP-2 and MMP-9 in cancer cells. Cancer Res; 70(5); 2095–104