Integrin-linked kinase (ILK) and estrogen receptor (ER)-alpha modulate cell migration. However, the crosstalk between ERalpha and ILK and the role of ILK in ERalpha-mediated cell migration remain unexplored. Here, we report that ILK participates in ERalpha signaling in breast cancer cells. We found that ILK binds ERalpha in vitro and in vivo through a LXXLL motif in ILK. Estrogen prevented ERalpha-ILK binding, resulting in phosphatidylinositol 3-kinase (PI3K)-dependent increase in ILK kinase activity. Furthermore, the regulation of ERalpha-ILK interaction was dependent on the PI3K pathway. Unexpectedly, transient knockdown or inhibition of ILK caused hyperphosphorylation of ERalpha Ser(118) in an extracellular signal-regulated kinase/mitogen-activated protein kinase pathway-dependent manner and an enhanced ERalpha recruitment to the target chromatin and gene expression, a process reversed by overexpression of ILK. Compatible with these interactions, estrogen regulated cell migration via the PI3K/ILK/AKT pathway with stable ILK overexpression hyperactivating cell migration. Thus, status of ILK signaling may be an important modifier of ER signaling in breast cancer cells and this pathway could be exploited for therapeutic intervention in breast cancer cells.