The HER family is composed of four receptor tyrosine kinases, which are frequently deregulated in several types of cancer. Activated HER receptors initiate intracellular signalling pathways by attracting to the plasma membrane a plethora of adaptor and signalling molecules. Although there are more than a dozen HER-interacting proteins that regulate signal transduction and have been extensively studied, recent proteomic studies have shown the existence of many novel but largely uncharacterized factors that may bind HER receptors. In this report, we describe a cell-based identification of several new HER2-binding proteins, including HAX1, YWHAZ, PELO and ACP1. Analysis of these factors showed that one of them, PELO, binds to active HER2 and epidermal growth factor receptor and thereby attenuates phosphatidylinositol 3-kinase (PI3K)/AKT signalling, likely through regulation of the recruitment of p85-PI3K to activated receptor. Functional characterization of PELO showed that it negatively regulates cell migration and metastasis in vivo. These results reveal that PELO is a novel regulator of HER-signalling and therefore is likely to have a role in inhibiting tumour progression and invasion.