Molecular Basis for Interaction between Icap1α PTB Domain and β1 Integrin

Abstract

Icap1␣ is a 200-amino acid protein that binds to theIntegrins are transmembrane heterodimeric receptors for extracellular matrix and cell surface proteins (1, 2). The binding of integrins to ligands in the extracellular matrix is linked to cell attachment and spreading, which in turn activates various cytosolic signal cascades to promote cell migration, survival, proliferation, and differentiation (3-5). The binding of integrins to their ligands requires integrins to be in an "activated" conformation (6). The regulation of the activation status of integrins and the post-ligand binding activation of various signaling cascades require the integrin cytoplasmic domains. Mutagenesis studies have shown that the cytoplasmic domain of ␤ subunit is important for cell adhesion and migration (7,8) and for localization of integrins to focal contacts (9, 10).Integrin ␤ subunit cytoplasmic domain interacts with several cytoskeletal proteins, including ␣-actinin, talin, paxillin, and filamin (11-16), which may localize integrins to the site of cell-extracellular matrix interaction. Signaling molecules, such as FAK, ILK-1 (17), RACK1 (18), and Shc (19,20), also bind to the integrin ␤ subunit cytoplasmic domains and may link integrins directly to the cytosolic kinase cascades. In addition to these proteins, which can bind to the cytoplasmic domains of several different ␤ subunits, there are proteins with binding specificity toward a particular ␤ integrin cytoplasmic domain. For example, Cytohesin-1 interacts specifically with the ␤ 2 integrin (21) and regulates the leukocyte-specific ␣ L ␤ 2 integrins (22). Similarly, ␤ 3 -endonexin binds to the ␤ 3 integrin (23) and may play a role in the affinity regulation of platelet integrin ␣ IIB ␤ 3 (24). Another of these proteins is Icap1␣, 1 which displays a restricted binding toward the ␤ 1 integrins (25).Although the precise function of Icap1␣ has not been established, its role in integrin-dependent cell adhesion was suggested from the finding that Icap1␣ undergoes an adhesion-dependent phosphorylation (25). Furthermore, expression of a mutant Icap1␣ with a T38D mutation interferes with cell spreading (26). The Icap1␣ binding site on the ␤ 1 integrin has been mapped to the COOH-terminal region of ␤ 1 integrin (25), which includes one of the two NPXY (or NXXY) motifs present in several integrin ␤ subunits. These NPXY motifs are important for the localization of integrins to focal contacts (10), integrin-mediated endocytosis (27), and affinity regulation of integrins (28). The functions of ␤ 1 and ␤ 3 integrins can also be regulated by tyrosine phosphorylation at the NPXY motifs (29 -31).Here, we examined the structural basis for the interaction between Icap1␣ and the integrin ␤ 1 cytoplasmic domain. Sequence homology and molecular modeling reveal that Icap1␣ has a PTB fold. The PTB domains bind their target peptides through extensive hydrogen bonding and packing interactions in which the target peptide is bound to a characteristic grove on the PTB domain. Furthermore, some PTB domains ...

Disruption of Focal Adhesions by Integrin Cytoplasmic Domain-associated Protein-1α

“…Indeed, sequence homology and molecular modeling favor the view that ICAP-1␣ is a phosphotyrosine binding domain protein. It was suggested that the interaction specificity with the ␤ 1A cytosolic tail was due to the interaction of Val-787 on the integrin and an hydrophobic pocket created by Leu-82 and Tyr-144 of ICAP-1␣ (25). This is fairly consistent with the lack of interaction of ICAP-1␣ with the ␤ 1D isoform that do not have a valine at this position.…”

“…ICAP-1 is expressed throughout development and also in adult tissues (24). ICAP-1␣ but not ICAP-1␤ interacts with the cytoplasmic tail of the ␤ 1A chain in a manner that depends on the conserved NPXY integrin motif (25). ICAP-1␣ contains a number of putative phosphorylation sites, including a phosphorylation motif for the CaMKII around threonine 38.…”

Integrin β cytoplasmic domain interactions with phosphotyrosine-binding domains: A structural prototype for diversity in integrin signaling

“…The interactions of ICAP-1␣ and talin are mediated by the binding of a PTB domain to an NPXYcontaining peptide sequence (6,7). Another group of proteins physically interact or colocalize with integrins, but their binding to the ␤ tails has not been established.…”

“…2 A-C), indicating that this region is important for integrin-PTB domain interactions. Most integrin ␤ tails have two NPXY motifs and the more C-terminal motif is implicated in binding to the ICAP-1␣ PTB domain (6). Consistent with a PTB domain-binding function for the second NPXY-like motif, the binding of Dab1 and Dab2 PTB domains to ␤3 tails was largely unaffected by a Tyr-747 to Ala mutation in the first NPXY motif of ␤3, but was inhibited by Tyr-759 to Ala mutations in the second NPXY-like motif (Fig.…”

Unraveling ICAP-1 function: Toward a new direction?

“…The N terminal moiety (1-60 aa) is rich in serine/threonine residues and contains several consensus sites for protein kinases such as PKC, PKA/PKG, PAKs and CaMKII (Ca2+/Calmodulin dependent protein kinase II). The C-terminal region (61-200 aa) has a phospho-tyrosine binding domain (PTB domain) that includes the binding site for the more distal NPxY motif of the β 1 integrin cytoplasmic tail (Chang et al, 2002). Using site directed mutagenesis and protein structure modeling, a number of key amino-acid residues on both β 1 integrin and ICAP-1 were proposed to be required for ICAP-1 integrin interaction.…”