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F Maina

Publications and source records attributed to F Maina.

14 recordsLinked to original sources
PubMed2001

Coupling Met to specific pathways results in distinct developmental outcomes.

Receptor tyrosine kinases (RTKs) mediate distinct biological responses by stimulating similar intracellular signaling pathways. Whether the specificity of the response is determined by qualitative or quantitative differences in signaling output is not known. We addressed this question in vivo by replacing the multifunctional docking sites of Met, the receptor for hepatocyte growth factor, with specific binding motifs for phosphatidylinositol-3 kinase, Src tyrosine kinase, or Grb2 (Met(2P), Met(2S), and Met(2G), respectively). All three mutants retained normal signaling through the multiadaptor Gab1, but differentially recruited specific effectors. While Met(2G) mice developed normally, Met(2P) and Met(2S) mice were loss-of-function mutants displaying different phenotypes and rescue of distinct tissues. These data indicate that RTK-mediated activation of specific signaling pathways is required to fulfill cell-specific functions in vivo.

Adaptor Proteins, Signal Transducing↗
PubMed2000

The branchial arches and HGF are growth-promoting and chemoattractant for cranial motor axons.

During development, cranial motor neurons extend their axons along distinct pathways into the periphery. For example, branchiomotor axons extend dorsally to leave the hindbrain via large dorsal exit points. They then grow in association with sensory ganglia, to their targets, the muscles of the branchial arches. We have investigated the possibility that pathway tissues might secrete diffusible chemorepellents or chemoattractants that guide cranial motor axons, using co-cultures in collagen gels. We found that explants of dorsal neural tube or hindbrain roof plate chemorepelled cranial motor axons, while explants of cranial sensory ganglia were weakly chemoattractive. Explants of branchial arch mesenchyme were strongly growth-promoting and chemoattractive for cranial motor axons. Enhanced and oriented axon outgrowth was also elicited by beads loaded with Hepatocyte Growth Factor (HGF); antibodies to this protein largely blocked the outgrowth and orientation effects of the branchial arch on motor axons. HGF was expressed in the branchial arches, whilst Met, which encodes an HGF receptor, was expressed by subpopulations of cranial motor neurons. Mice with targetted disruptions of HGF or Met showed defects in the navigation of hypoglossal motor axons into the branchial region. Branchial arch tissue may thus act as a target-derived factor that guides motor axons during development. This influence is likely to be mediated partly by Hepatocyte Growth Factor, although a component of branchial arch-mediated growth promotion and chemoattraction was not blocked by anti-HGF antibodies.

Animals↗
PubMed2000

Met signaling mutants as tools for developmental studies.

The Met receptor is widely expressed in embryonic and adult epithelial tissues; its ligand (hepatocyte growth factor/scatter factor, HGF/SF) is expressed in the mesenchymal component of various organs. The generation of hgf and met null mice has revealed an essential role for this ligand-receptor pair in the development of the placenta, liver, and limb muscles. However the early lethality of the null mutants has precluded analysis of Met function in late development. To extend the possible observation period, we generated mutant metalleles of different severity. This was done by impairing the ability of the receptor to transduce the HGF/SF signal, via mutation of consensus sequences in the multifunctional docking site present in the C-terminal tail of the receptor. Mice expressing a Met mutant still active as a kinase, but unable to recruit its effectors, died in mid-gestation with the same phenotype as the metknockout, proving the importance of phosphotyrosine-SH2 interactions in vivo. Mice expressing a Met receptor with partial loss of signaling function survived until birth and revealed novel aspects of HGF/SF-Met function during muscle development.

Animals↗
PubMed1999

Hepatocyte growth factor, a versatile signal for developing neurons.

Here we summarize recent findings on the biology of hepatocyte growth factor (HGF, also known as scatter factor), focusing on its effects on developing neurons. HGF is both a chemoattractant and a survival factor for embryonic motor neurons. In addition, sensory and sympathetic neurons and their precursors respond to HGF with increased differentiation, survival and axonal outgrowth. The broad spectrum of HGF activities and its observed synergy with other neurotrophic factors suggest that the major role of HGF is to potentiate the response of developing neurons to specific signals.

Animals↗
PubMed1999

Hepatocyte growth factor/scatter factor-MET signaling in neural crest-derived melanocyte development.

The mechanisms governing development of neural crest-derived melanocytes, and how alterations in these pathways lead to hypopigmentation disorders, are not completely understood. Hepatocyte growth factor/scatter factor (HGF/SF) signaling through the tyrosine-kinase receptor, MET, is capable of promoting the proliferation, increasing the motility, and maintaining high tyrosinase activity and melanin synthesis of melanocytes in vitro. In addition, transgenic mice that ubiquitously overexpress HGF/SF demonstrate hyperpigmentation in the skin and leptomenigenes and develop melanomas. To investigate whether HGF/ SF-MET signaling is involved in the development of neural crest-derived melanocytes, transgenic embryos, ubiquitously overexpressing HGF/SF, were analyzed. In HGF/SF transgenic embryos, the distribution of melanoblasts along the characteristic migratory pathway was not affected. However, additional ectopically localized melanoblasts were also observed in the dorsal root ganglia and neural tube, as early as 11.5 days post coitus (p.c.). We utilized an in vitro neural crest culture assay to further explore the role of HGF/SF-MET signaling in neural crest development. HGF/SF added to neural crest cultures increased melanoblast number, permitted differentiation into pigmented melanocytes, promoted melanoblast survival, and could replace mast-cell growth factor/Steel factor (MGF) in explant cultures. To examine whether HGF/SF-MET signaling is required for the proper development of melanocytes, embryos with a targeted Met null mutation (Met-/-) were analysed. In Met-/- embryos, melanoblast number and location were not overtly affected up to 14 days p.c. These results demonstrate that HGF/SF-MET signaling influences, but is not required for, the initial development of neural crest-derived melanocytes in vivo and in vitro.

Animals↗
PubMed1998

Multiple roles for hepatocyte growth factor in sympathetic neuron development.

We have studied the role of hepatocyte growth factor (HGF)/Met signaling in the development of sympathetic neuroblasts and neurons. Anti-HGF antibodies reduced the number of sympathetic neuroblasts that differentiated into neurons, but neither anti-HGF antibodies nor HGF affected neuroblast proliferation. Anti-HGF antibodies also reduced the survival of neuroblasts but not sympathetic neurons. HGF greatly enhanced the neurite outgrowth of NGF-dependent sympathetic neurons throughout development. These in vitro effects of anti-HGF antibodies and HGF were abolished by a disabling mutation of Met, the HGF receptor tyrosine kinase. The Met mutation also increased sympathetic neuroblast apoptosis in vivo. Because Met and HGF are expressed in sympathetic ganglia throughout development, it is possible that the multiple effects of HGF/Met signaling on sympathetic neuroblasts and neurons occur in part by an autocrine mechanism.

Animals↗
PubMed1997

Met receptor signaling is required for sensory nerve development and HGF promotes axonal growth and survival of sensory neurons.

The development of the nervous system is a dynamic process during which factors act in an instructive fashion to direct the differentiation and survival of neurons, and to induce axonal outgrowth, guidance to, and terminal branching within the target tissue. Here we report that mice expressing signaling mutants of the hepatocyte growth factor (HGF) receptor, the Met tyrosine kinase, show a striking reduction of sensory nerves innervating the skin of the limbs and thorax, implicating the HGF/Met system in sensory neuron development. Using in vitro assays, we find that HGF cooperates with nerve growth factor (NGF) to enhance axonal outgrowth from cultured dorsal root ganglion (DRG) neurons. HGF also enhances the neurotrophic activities of NGF in vitro, and Met receptor signaling is required for the survival of a proportion of DRG neurons in vivo. This synergism is specific for NGF but not for the related neurotrophins BDNF and NT3. By using a mild signaling mutant of Met, we have demonstrated previously that Met requires signaling via the adapter molecule Grb2 to induce proliferation of myoblasts. In contrast, the actions of HGF on sensory neurons are mediated by Met effectors distinct from Grb2. Our findings demonstrate a requirement for Met signaling in neurons during development.

Animals↗
PubMed1996

Uncoupling of Grb2 from the Met receptor in vivo reveals complex roles in muscle development.

Hepatocyte growth factor (HGF) and its receptor, the Met tyrosine kinase, are determinants of placenta, liver, and muscle development. Here, we show that Met function in vivo requires signaling via two carboxy-terminal tyrosines. Mutation of both residues in the mouse genome caused embryonal death, with placenta, liver, and limb muscle defects, mimicking the phenotype of met null mutants. In contrast, disrupting the consensus for Grb2 binding allowed development to proceed to term without affecting placenta and liver but caused a striking reduction in limb muscle coupled to a generalized deficit of secondary fibers. These data show that the requirements for Met signaling vary depending on the tissue and reveal a novel role for HGF/ Met in late myogenesis.

Adaptor Proteins, Signal Transducing↗
PubMed1996

Specific uncoupling of GRB2 from the Met receptor. Differential effects on transformation and motility.

The biological effects of hepatocyte growth factor/scatter factor are mediated by autophosphorylation of its receptor, the Met tyrosine kinase, on two carboxyl-terminal tyrosines. These phosphotyrosines (Y1349VHVNATY1356VNV) are multifunctional docking sites for several effectors. Grb2, the adaptor for the Ras guanyl-nucleotide exchanger SOS, binds to Tyr1356 in the YVNV motif. By site-directed mutagenesis we either abrogated or duplicated the Grb2 consensus, without interfering with the other effectors. Loss of the link with Grb2 severely impaired transformation. The same mutation, however, had no effect on the "scattering" response, indicating that the level of signal which can be reached by Grb2-independent routes is permissive for motility. Duplication of the Grb2 binding site enhanced transformation and left motility unchanged. Thus, two Met-mediated biological responses, motility and growth, can be dissociated on the basis of their differential requirement for a direct link with Ras.

3T3 Cells↗
PubMed1994

A multifunctional docking site mediates signaling and transformation by the hepatocyte growth factor/scatter factor receptor family.

Signaling by tyrosine kinase receptors is mediated by selective interactions between individual Src homology 2 (SH2) domains of cytoplasmic effectors and specific phosphotyrosine residues in the activated receptor. Here, we report the existence in the hepatocyte growth factor/scatter factor (HGF/SF) receptor of a multifunctional docking site made of the tandemly arranged degenerate sequence YVH/NV. Phosphorylation of this site mediates intermediate- to high-affinity interactions with multiple SH2-containing signal transducers, including phosphatidylinositol 3-kinase, phospholipase C gamma, pp60c-src, and the GRB-2-Sos complex. Mutation of the two tyrosines results in loss of biological function, as shown by abrogation of the transforming activity in the oncogenic counterpart of the receptor. The same bidentate motif is conserved in the evolutionarily related receptors Sea and Ron, suggesting that in all members of the HGF/SF receptor family, signal transduction is channeled through a multifunctional binding site.

Adaptor Proteins, Signal Transducing↗
PubMed1994

Hepatocyte growth factor and its receptor, the tyrosine kinase encoded by the c-MET proto-oncogene.

HGF is secreted by mesenchymal cells and regulates motogenesis, mitogenesis, and morphogenesis of epithelial and endothelial cells. HGF is a heterodimer of two glycosylated chains, alpha and beta, bound together by a disulfide bond. The molecule is synthesized as single chain precursor devoid of biological activity (pro-HGF). The critical step in pro-HGF activation is a proteolytic cleavage generating the two chain form. This step occurs in the extracellular environment, and is catalyzed by urokinase. Two alternative transcripts originate two HGF variants. One bears a deletion of five amino acids in the alpha chain, and has the same properties of the full-size protein. The other one contains only the first portion of the alpha chain (two kringle HGF). Two kringle HGF binds the HGF receptor, triggers its tyrosine kinase activity and behaves as a partial agonist, inducing motogenesis but not mitogenesis in target cells. The HGF receptor is the tyrosine kinase encoded by the c-MET pro-oncogene, a tyrosine kinase receptor. This molecule is an heterodimer of an extracellular alpha chain disulfide linked to a transmembrane beta chain. The cytoplasmic portion of the beta chain contains the catalytic domain and critical sites for the regulation of its kinase activity. In the C-terminal tail, a bidentate motif containing two tyrosines associates the transducers responsible for HGF signalling.

Amino Acid Sequence↗
PubMed1993

A novel recognition motif for phosphatidylinositol 3-kinase binding mediates its association with the hepatocyte growth factor/scatter factor receptor.

The pleiotropic effects (mitogenesis, motogenesis, and morphogenesis) elicited by hepatocyte growth factor/scatter factor (HGF/SF) are mediated by the activation of the tyrosine kinase receptor encoded by the MET proto-oncogene. Following autophosphorylation, the receptor associates with the p85/110 phosphatidylinositol (PI) 3-kinase complex in vivo and in vitro. By a combination of two complementary approaches, competition with synthetic phosphopeptides and association with Tyr-Phe receptor mutants, we have identified Y-1349 and Y-1356 in the HGF/SF receptor as the binding sites for PI 3-kinase. Y-1349VHV and Y-1356VNV do not conform to the canonical consensus sequence YXXM for PI 3-kinase binding and thus define YVXV as a novel recognition motif. Y-1349 and Y-1356 are located within the C-terminal portion of the HGF/SF receptor and are phosphorylation sites. The affinity of the N- and C-terminal src homology region 2 (SH2) domains of p85 for the phosphopeptides including Y-1349 and Y-1356 is 2 orders of magnitude lower than that measured for Y-751 in the platelet-derived growth factor receptor binding site. However, the closely spaced duplication of the novel recognition motif in the native HGF/SF receptor may allow binding with both SH2 domains of p85, thus generating an efficient docking site for PI 3-kinase. In agreement with this model, we have observed that a phosphopeptide including both Y-1349 and Y-1356 activates PI 3-kinase in vitro.

Amino Acid Sequence↗
PubMed1992

Autophosphorylation promotes complex formation of recombinant hepatocyte growth factor receptor with cytoplasmic effectors containing SH2 domains.

Hepatocyte growth factor (HGF), also known as scatter factor (SF), is a polypeptide which induces motility and/or mitogenesis in epithelial cells. The receptor for HGF/SF, p190MET, is a two-chain transmembrane tyrosine kinase encoded by the MET proto-oncogene. To identify the cytoplasmic effectors involved in signal transduction we have produced the human HGF/SF receptor in insect cells (Sf9) by means of a recombinant baculovirus. Two 170-kDa forms of the receptor were synthesized in Sf9 cells: the uncleaved single-chain precursor (which is by far the more abundant) and the proteolytically processed two-chain molecule. Both receptor species are phosphorylated on tyrosine in vivo and are active kinases in vitro. The recombinant receptor binds and phosphorylates in vitro four known cytoplasmic transducers containing src homology region 2 (SH2) domains: the 85-kDa subunit of phosphatidylinositol 3-kinase (Pl 3-kinase), rasGAP, phospholipase-C gamma (PLC-gamma), and p59Fyn, a tyrosine kinase of the src family. In all cases the association is strictly dependent on tyrosine phosphorylation of the receptor, indicating that it occurs via specific interaction with the SH2 domains. These results show that the HGF/SF receptor has the sequence requirements for binding a spectrum of cytoplasmic transducers whose different combinations in target cells may result in the observed pleiotropic biological response.

Animals↗