Ena/VASP proteins are actin-associated proteins involved in a range of processes dependent on cytoskeleton remodeling and cell polarity such as axon guidance and lamellipodial and filopodial dynamics in migrating cells. ENAH induces the formation of F-actin rich outgrowths in fibroblasts. Acts synergistically with BAIAP2-alpha and downstream of NTN1 to promote filipodia formation (By similarity).
The intracellular targeting of Ena/VASP family members is achieved via the interaction of their EVH1 domain with FPPPP sequence motifs found in a variety of cytoskeletal proteins, including lamellipodin, vinculin, and zyxin. Here we show that the LIM3 domain of Tes, which lacks the FPPPP motif, binds to the EVH1 domain of Mena, but not to those of VASP or Evl. The structure of the LIM3:EVH1 complex reveals that Tes occludes the FPPPP-binding site and competes with FPPPP-containing proteins for EVH1 binding. Structure-based gain-of-function experiments define the molecular basis for the specificity of the Tes-Mena interaction. Consistent with in vitro observations, the LIM3 domain displaces Mena, but not VASP, from the leading edge and focal adhesions. It also regulates cell migration through a Mena-dependent mechanism. Our observations identify Tes as an atypical EVH1 binding partner and a regulator specific to a single Ena/VASP family member.
BACKGROUND: The Rho GTPases Rho, Rac, and Cdc42 regulate the organization of the actin cytoskeleton by interacting with multiple, distinct downstream effector proteins. Cdc42 controls the formation of actin bundle-containing filopodia at the cellular periphery. The molecular mechanism for this remains as yet unclear. RESULTS: We report here that Cdc42 interacts with IRSp53/BAP2 alpha, an SH3 domain-containing scaffold protein, at a partial CRIB motif and that an N-terminal fragment of IRSp53 binds, via an intramolecular interaction, to the CRIB motif-containing central region. Overexpression of IRSp53 in fibroblasts leads to the formation of filopodia, and both this and Cdc42-induced filopodia are inhibited by expression of the N-terminal IRSp53 fragment. Using affinity chromatography, we have identified Mena, an Ena/VASP family member, as interacting with the SH3 domain of IRSp53. Mena and IRSp53 act synergistically to promote filopodia formation. CONCLUSION: We conclude that the interaction of Cdc42 with the partial CRIB motif of IRSp53 relieves an intramolecular, autoinhibitory interaction with the N terminus, allowing the recruitment of Mena to the IRSp53 SH3 domain. This IRSp53:Mena complex initiates actin filament assembly into filopodia.
Interacting selectively and non-covalently with a SH3 domain (Src homology 3) of a protein, small protein modules containing approximately 50 amino acid residues found in a great variety of intracellular or membrane-associated proteins.
Interacting selectively and non-covalently with a WW domain of a protein, a small module composed of 40 amino acids and plays a role in mediating protein-protein interactions via proline-rich regions.
Evidence
1:
Inferred from Physical InteractionUniProtKB
The neuronal protein FE65 functions in brain development and amyloid precursor protein (APP) signaling through its interaction with the mammalian enabled (Mena) protein and APP, respectively. The recognition of short polyproline sequences in Mena by the FE65 WW domain has a central role in axon guidance and neuronal positioning in the developing brain. We have determined the crystal structures of the human FE65 WW domain (residues 253-289) in the apo form and bound to the peptides PPPPPPLPP and PPPPPPPPPL, which correspond to human Mena residues 313-321 and 347-356, respectively. The FE65 WW domain contains two parallel ligand-binding grooves, XP (formed by residues Y269 and W280) and XP2 (formed by Y269 and W271). Both Mena peptides adopt a polyproline helical II conformation and bind to the WW domain in a forward (N-C) orientation through selection of the PPPPP motif by the XP and XP2 grooves. This mode of ligand recognition is strikingly similar to polyproline interaction with SH3 domains. Importantly, comparison of the FE65 WW structures in the apo and liganded forms shows that the XP2 groove is formed by an induced-fit mechanism that involves movements of the W271 and Y269 side-chains upon ligand binding. These structures elucidate the molecular determinants underlying polyproline ligand selection by the FE65 WW domain and provide a framework for the design of small molecules that would interfere with FE65 WW-ligand interaction and modulate neuronal development and APP signaling.
A reference proteome is a set of protein sequences derived from a complete proteome which constitutes a defined standard for a particular user community. Reference proteomes are manually defined according to a number of criteria. They cover the proteomes of well- studied model organisms and other proteomes of interest for biomedical and biotechnological research. Reference proteomes have been selected to provide broad coverage of the tree of life, and constitute a representative cross-section of the taxonomic diversity to be found within UniProtKB.
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