The Bcr Kinase Downregulates Ras Signaling by Phosphorylating AF-6 and Binding to Its PDZ Domain

Abstract

The protein kinase Bcr is a negative regulator of cell proliferation and oncogenic transformation. We identified Bcr as a ligand for the PDZ domain of the cell junction and Ras-interacting protein AF-6. The Bcr kinase phosphorylates AF-6, which subsequently allows efficient binding of Bcr to AF-6, showing that the Bcr kinase is a regulator of the PDZ domain-ligand interaction. Bcr and AF-6 colocalize in epithelial cells at the plasma membrane. In addition, Bcr, AF-6, and Ras form a trimeric complex. Bcr increases the affinity of AF-6 to Ras, and a mutant of AF-6 that lacks a specific phosphorylation site for Bcr shows a reduced binding to Ras. Wild-type Bcr, but not Bcr mutants defective in binding to AF-6, interferes with the Ras-dependent stimulation of the Raf/MEK/ERK pathway. Since AF-6 binds to Bcr via its PDZ domain and to Ras via its Ras-binding domain, we propose that AF-6 functions as a scaffold-like protein that links Bcr and Ras to cellular junctions. We suggest that this trimeric complex is involved in downregulation of Ras-mediated signaling at sites of cell-cell contact to maintain cells in a nonproliferating state.

Abr and Bcr, Two Homologous Rac GTPase-Activating Proteins, Control Multiple Cellular Functions of Murine Macrophages

“…Recent studies have suggested that Bcr localizes to the plasma membrane at the cellular junctions of epithelial cells (34,42). Therefore, possible transient changes in Bcr and Abr localization could occur depending on the cell type and stimulus.…”

Essential Role of p38γ in K-Ras Transformation Independent of Phosphorylation

“…This motif has been shown to be important for its subcellular localizations and/or its interactions with other PDZ domain-containing proteins to maintain certain structures of cytoskeleton, a process that is important for Ras transformation (51,52). Moreover, these interactions are also regulated by protein phosphorylations and dephosphorylations (51,53).…”

Loss of polarity protein AF6 promotes pancreatic cancer metastasis by inducing Snail expression

“…3), and was originally identified as a fusion partner of the ALL-1 gene involved in acute lymphoblastic leukaemia 4 . AF6 is a multi-domain protein, consisting (from the N to C terminus) of two Ras-associating domains, a forkhead-associated domain, a dilute domain, a PDZ (PSD95/Dlg1/ZO-1) domain and three proline-rich regions (Pro) [5][6][7] . In rodents and humans, AF6 is expressed in almost all epithelial tissues 8 mainly with two isoforms: a long isoform containing the F-actin-binding domain (FA) and a short isoform lacking the FA domain; these versions are referred to as l-and s-afadin, respectively.…”