Jinhua Wu, PhD
Office Phone: 215-728-2867
1. Structural and biochemical characterization of the functional units of MRL proteins.
The enhanced motility of malignant tumor cells supports their invasion of surrounding tissue. Preventing the spread of cancer cells has become an important therapeutic approach for treating a variety of cancers. Recently, cell motility and actin dynamics have been found to associate with a group of adapter signaling proteins, designated the MRL family (based on family members MIG-10/RIAM/Lamellipodin). MRL proteins contain an RA-PH structural unit. This structural unit is responsible for both Ras GTPase binding and membrane localization of MRL. In part based on these interaction properties, these proteins are recognized as an important convergence point that links upstream signaling with actin dynamics.
RIAM and lamellipodin, the mammalian members of the MRL group, were shown to regulate cell migration by
interacting with the Ena/VASP proteins. We are interested in investigating the structural basis of the recruitment of MRL proteins to the Ena/VASP actin-binding proteins. The EVH1 domain of Ena/VASP binds specifically to the FP4 motifs in RIAM and lamellipodin. Previous study reveals that the RA-PH structural unit of MRL proteins may oligomerize through the amino terminal coiled-coil region. Our ongoing work indicates that the MRL proteins may function in clustering actin filaments that are bound to the Ena/VASP tetramer, thereby resulting in polymerization of actin filaments at the leading edge of the lamellipodium.Top
2. Molecular mechanisms governing the activation of integrin mediated by RIAM
The small GTPase Rap1 induces integrin activation via an inside-out signaling pathway mediated by the Rap1-interacting adaptor molecule (RIAM). Blocking this pathway may suppress tumour metastasis and other diseases that are related to hyperactive integrins. However, the molecular basis for the specific recognition of RIAM by Rap1 remains largely unknown. Herein we present the crystal structure of an active, GTP-bound GTPase domain of Rap1 in complex with the Ras association (RA)-pleckstrin homology (PH) structural module of RIAM at 1.65 Å. The structure reveals that the recognition of RIAM by Rap1 is governed by side-chain interactions. Several side chains are critical in determining specificity of this recognition, particularly the Lys31 residue in Rap1 that is oppositely charged compared with the Glu31/Asp31 residue in other Ras GTPases. Lys31 forms a salt bridge with RIAM residue Glu212, making it the key specificity determinant of the interaction. We also show that disruption of these interactions results in reduction of Rap1:RIAM association, leading to a loss of co-clustering and cell adhesion. Our findings elucidate the molecular mechanism by which RIAM mediates Rap1-induced integrin activation. The crystal structure also offers new insight into the structural basis for the specific recruitment of RA-PH module-containing effector proteins by their small GTPase partners.Top