Cytoplasmic ACK1 interaction with multiple receptor tyrosine kinases is mediated by Grb2: an analysis of ACK1 effects on Axl signaling

Ack1 (activated Cdc42-associated kinase 1), a cytoplsmic tyrosine kinase, is implicated in metastatic behavior, cell spreading and migration, and epidermal growth factor receptor (EGFR) signaling. The function of Ack1 in the regulation of Receptor Tyrosine Kinases requires a C-terminal region that demonstrates a significant homology to the EGFR binding domain of MIG6. In this study, we have identified additional Receptor Tyrosine Kinases, including Axl, leukocyte tyrosine kinase, and anaplastic lymphoma kinase, that can bind to the Ack1/MIG6 homology region. Unlike the interaction between MIG6 and EGFR, our data suggest that these Receptor Tyrosine Kinases require the adaptor protein Grb2 for efficient binding, which interacts with highly conserved proline-rich regions that are conserved between Ack1 and MIG6. We have focused on Axl and compared how Ack1/Axl differs from the Ack1/EGFR axis by investigating effects of knockdown of endogenous Ack1. Although EGFR activation promotes Ack1 turnover, Axl activation by GAS6 does not; interestingly, the reciprocal down-regulation of GAS6-stimulated Axl is blocked by removing Ack1. Thus, Ack1 functions in part to control Axl receptor levels. Silencing of Ack1 also leads to diminished ruffling and migration in DU145 and COS7 cells upon GAS6-Axl signaling. The ability of Ack1 to modulate Axl and perhaps anaplastic lymphoma kinase (altered in anaplastic large cell lymphomas) might explain why Ack1 can promote metastatic and transformed behavior in a number of cancers.