Direct interactions with the integrin ß1 cytoplasmic tail activate the Abl2/Arg kinase.

Integrins are heterodimeric α/ß extracellular matrix adhesion receptors that couple physically to the actin cytoskeleton and regulate kinase signaling pathways to control cytoskeletal remodeling and adhesion complex formation and disassembly. ß1 integrins signal through the Abl2/Arg (Abl-related gene) nonreceptor tyrosine kinase to control fibroblast cell motility, neuronal dendrite morphogenesis and stability, and cancer cell invasiveness, but the molecular mechanisms by which integrin ß1 activates Arg are unknown. We report here that the Arg kinase domain interacts directly with a lysine-rich membrane-proximal segment in the integrin ß1 cytoplasmic tail, that Arg phosphorylates the membrane-proximal Tyr-783 in the ß1 tail, and that the Arg Src homology domain then engages this phosphorylated region in the tail. We show that these interactions mediate direct binding between integrin ß1 and Arg in vitro and in cells and activate Arg kinase activity. These findings provide a model for understanding how ß1-containing integrins interact with and activate Abl family kinases.

Abl2/Abl-related gene stabilizes actin filaments, stimulates actin branching by actin-related protein 2/3 complex, and promotes actin filament severing by cofilin.

Both Arp2/3 complex and the Abl2/Arg nonreceptor tyrosine kinase are essential to form and maintain diverse actin-based structures in cells, including cell edge protrusions in fibroblasts and cancer cells and dendritic spines in neurons. The ability of Arg to promote cell edge protrusions in fibroblasts does not absolutely require kinase activity, raising the question of how Arg might modulate actin assembly and turnover in the absence of kinase function. Arg has two distinct actin-binding domains and interacts physically and functionally with cortactin, an activator of the Arp2/3 complex. However, it was not known whether and how Arg influences actin filament stability, actin branch formation, or cofilin-mediated actin severing or how cortactin influences these reactions of Arg with actin. Arg or cortactin bound to actin filaments stabilizes them from depolymerization. Low concentrations of Arg and cortactin cooperate to stabilize filaments by slowing depolymerization. Arg stimulates formation of actin filament branches by Arp2/3 complex and cortactin. An Arg mutant lacking the C-terminal calponin homology actin-binding domain stimulates actin branch formation by the Arp2/3 complex, indicative of autoinhibition. ArgΔCH can stimulate the Arp2/3 complex even in the absence of cortactin. Arg greatly potentiates cofilin severing of actin filaments, and cortactin attenuates this enhanced severing. The ability of Arg to stabilize filaments, promote branching, and increase severing requires the internal (I/L)WEQ actin-binding domain. These activities likely underlie important roles that Arg plays in the formation, dynamics, and stability of actin-based cellular structures.

Long-range architecture in a viral RNA genome.

We have developed a model for the secondary structure of the 1058-nucleotide plus-strand RNA genome of the icosahedral satellite tobacco mosaic virus (STMV) using nucleotide-resolution SHAPE chemical probing of the viral RNA isolated from virions and within the virion, perturbation of interactions distant in the primary sequence, and atomic force microscopy. These data are consistent with long-range base pairing interactions and a three-domain genome architecture. The compact domains of the STMV RNA have dimensions of 10-45 nm. Each of the three domains corresponds to a specific functional component of the virus: The central domain corresponds to the coding sequence of the single (capsid) protein encoded by the virus, whereas the 5' and 3' untranslated domains span signals essential for translation and replication, respectively. This three-domain architecture is compatible with interactions between the capsid protein and short RNA helices previously visualized by crystallography. STMV is among the simplest of the icosahedral viruses but, nonetheless, has an RNA genome with a complex higher-order structure that likely reflects high information content and an evolutionary relationship between RNA domain structure and essential replicative functions.

ß1 integrin regulates Arg to promote invadopodial maturation and matrix degradation.

ß1 integrin has been shown to promote metastasis in a number of tumor models, including breast, ovarian, pancreatic, and skin cancer; however, the mechanism by which it does so is poorly understood. Invasive membrane protrusions called invadopodia are believed to facilitate extracellular matrix degradation and intravasation during metastasis. Previous work showed that ß1 integrin localizes to invadopodia, but its role in regulating invadopodial function has not been well characterized. We find that ß1 integrin is required for the formation of mature, degradation-competent invadopodia in both two- and three-dimensional matrices but is dispensable for invadopodium precursor formation in metastatic human breast cancer cells. ß1 integrin is activated during invadopodium precursor maturation, and forced ß1 integrin activation enhances the rate of invadopodial matrix proteolysis. Furthermore, ß1 integrin interacts with the tyrosine kinase Arg and stimulates Arg-dependent phosphorylation of cortactin on tyrosine 421. Silencing ß1 integrin with small interfering RNA completely abrogates Arg-dependent cortactin phosphorylation and cofilin-dependent barbed-end formation at invadopodia, leading to a significant decrease in the number and stability of mature invadopodia. These results describe a fundamental role for ß1 integrin in controlling actin polymerization-dependent invadopodial maturation and matrix degradation in metastatic tumor cells.

Integrin ß1 signals through Arg to regulate postnatal dendritic arborization, synapse density, and behavior.

Integrins are heterodimeric extracellular matrix receptors that are essential for the proper development of the vertebrate nervous system. We report here that selective loss of integrin ß1 in excitatory neurons leads to reductions in the size and complexity of hippocampal dendritic arbors, hippocampal synapse loss, impaired hippocampus-dependent learning, and exaggerated psychomotor sensitivity to cocaine in mice. Our biochemical and genetic experiments demonstrate that the intracellular tail of integrin ß1 binds directly to Arg kinase and that this interaction stimulates activity of the Arg substrate p190RhoGAP, an inactivator of the RhoA GTPase. Moreover, genetic manipulations that reduce integrin ß1 signaling through Arg recapitulate the integrin ß1 knock-out phenotype in a gene dose-sensitive manner. Together, these results describe a novel integrin ß1-Arg-p190RhoGAP pathway that regulates dendritic arbor size, promotes synapse maintenance, supports proper hippocampal function, and mitigates the behavioral consequences of cocaine exposure.