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M Grumet

Publications and source records attributed to M Grumet.

At least 37 records · Page 2Linked to original sources
PubMed1998

Multi-ligand interactions with receptor-like protein tyrosine phosphatase beta: implications for intercellular signaling.

Receptor-like protein tyrosine phosphatase beta (RPTP beta) shows structural and functional similarity to cell adhesion molecules (CAMs). It binds to several neuronal CAMs and extracellular matrix (ECM) proteins that combine to form cell-recognition complexes. Here, the authors discuss the implications of such complexes for intercellular signaling, and the regulation of RPTP activity by cell-cell and cell-ECM contact.

Animals↗
PubMed1997

Cell adhesion molecules regulate guidance of dorsal root ganglion axons in the marginal zone and their invasion into the mantle layer of embryonic spinal cord.

In order to elucidate the mechanisms regulating the projections of dorsal root ganglion (DRG) axons in the dorsal funiculus and invasion into target regions in the mantle layer (prospective gray matter) of the spinal cord, we examined the interactions between DRG axons and spinal cord. DRG neurons were dissociated from chick embryos and cultured for 1-2 days on cryostat sections of the spinal cord at embryonic day 5 (E5) or at E9. E5 and E9 DRG neurons extended neurites onto both marginal zone (prospective white matter) and mantle layer (prospective gray matter) of the spinal cord, suggesting that both of these regions are permissive for neurite growth. When E5 DRG neurites approached cryosections of E5 spinal cord from outside, most of them ran in the marginal zone without invading the mantle layer. In contrast, about half of E9 DRG neurites entered the mantle layer after crossing the marginal zone of E9 spinal cord. These growth patterns of DRG neurites on spinal marginal zone and mantle layer are similar to the pathway formation of DRG axons at comparable stages in vivo; DRG axons run exclusively in the prospective dorsal funiculus before E6, and enter the mantle layer (prospective dorsal horn) to reach the target regions by E9. Perturbation of functions of Ng-CAM, Nr-CAM, and axonin-1/SC2 by adding the specific antibodies in the culture medium increased the ratio of DRG neurites entering the mantle layer of E5 spinal cord, suggesting that these cell adhesion molecules are involved in keeping DRG neurites in the marginal zone. Taken together with the expression of Ng-CAM, Nr-CAM, and axonin-1/SC2, these CAMs on DRG axons may regulate the guidance of these axons in the marginal zone before E6, and the subsequent decrease in the relative levels of these CAMs might allow DRG axons to invade the target mantle layer.

Animals↗
PubMed1997

Identification of a novel contactin-associated transmembrane receptor with multiple domains implicated in protein-protein interactions.

Receptor protein tyrosine phosphatase beta (RPTPbeta) expressed on the surface of glial cells binds to the glycosylphosphatidylinositol (GPI)-anchored recognition molecule contactin on neuronal cells leading to neurite outgrowth. We describe the cloning of a novel contactin-associated transmembrane receptor (p190/Caspr) containing a mosaic of domains implicated in protein-protein interactions. The extracellular domain of Caspr contains a neurophilin/coagulation factor homology domain, a region related to fibrinogen beta/gamma, epidermal growth factor-like repeats, neurexin motifs as well as unique PGY repeats found in a molluscan adhesive protein. The cytoplasmic domain of Caspr contains a proline-rich sequence capable of binding to a subclass of SH3 domains of signaling molecules. Caspr and contactin exist as a complex in rat brain and are bound to each other by means of lateral (cis) interactions in the plasma membrane. We propose that Caspr may function as a signaling component of contactin, enabling recruitment and activation of intracellular signaling pathways in neurons. The binding of RPTPbeta to the contactin-Caspr complex could provide a mechanism for cell-cell communication between glial cells and neurons during development.

Amino Acid Sequence↗
PubMed1997

Induction of neurite outgrowth through contactin and Nr-CAM by extracellular regions of glial receptor tyrosine phosphatase beta.

Receptor protein tyrosine phosphatase beta (RPTPbeta) is expressed as soluble and receptor forms with common extracellular regions consisting of a carbonic anhydrase domain (C), a fibronectin type III repeat (F), and a unique region called S. We showed previously that a recombinant Fc fusion protein with the C domain (beta C) binds to contactin and supports neuronal adhesion and neurite growth. As a substrate, betaCFS was less effective in supporting cell adhesion, but it was a more effective promoter of neurite outgrowth than betaCF. betaS had no effect by itself, but it potentiated neurite growth when mixed with betaCF. Neurite outgrowth induced by betaCFS was inhibited by antibodies against Nr-CAM and contactin, and these cell adhesion molecules formed a complex that bound betaCFS. NIH-3T3 cells transfected to express betaCFS on their surfaces induced neuronal differentiation in culture. These results suggest that binding of glial RPTPbeta to the contactin/Nr-CAM complex is important for neurite growth and neuronal differentiation.

Animals↗
PubMed1997

Nr-CAM: a cell adhesion molecule with ligand and receptor functions.

Nr-CAM Ng-CAM-related cell adhesion molecule) is expressed only in the nervous system on a subset of neurons and non-neuronal cells, including floor-plate cells in the spinal cord and Schwann cells. It is a member of the immunoglobulin (Ig) superfamily and can bind by a homophilic mechanism but its heterophilic interactions may be of greater biological significance. Nr-CAM functions as a neuronal receptor for neurite-growth-promoting stimuli provided by contactin/F11, neurofascin, and RPTPbeta. In addition, by binding to neuronal receptors such as axonin-1, it can modulate axonal guidance. Nr-CAM can also interact laterally with contactin/F11 within the plasma membrane in the form of a complex that may transmit signals to regulate axonal growth. This review summarizes the structure and expression of Nr-CAM and discusses its potential functions as a ligand and as a receptor during neural development.

Animals↗
PubMed1997

Tenascin-C expression in the cyst wall and fluid of human brain tumors correlates with angiogenesis.

OBJECTIVE: Tenascin-C (TN) is an extracellular matrix glycoprotein with a characteristic six-armed structure. The aim of this study was to determine whether the concentration of TN in the cyst fluid of brain tumors can be used as a marker for angiogenesis and glioma grade. METHODS: We investigated the expression of TN in the cyst wall and cyst fluid of human brain tumors by immunohistochemistry, immunoprecipitation, and immunoblotting. The tumors included 12 astrocytomas (5 glioblastoma multiforme tumors, 1 anaplastic astrocytoma, 1 low-grade astrocytoma, 4 juvenile pilocytic astrocytomas, and 1 mixed glioma), 2 dysembryoplastic neuroepithelial tumors, 3 craniopharyngiomas, 2 ependymomas, 2 metastatic carcinomas, 3 arachnoid cysts, 1 glial ependymal cyst, and 1 inflammatory cyst. RESULTS: We detected no expression of TN in the cyst fluids of the ependymomas, craniopharyngiomas, and nonpilocytic low-grade astrocytoma. By contrast, TN was detected in the cyst fluids of all the other tumors. Results of quantitative immunoblotting using a PhosphorImager unit (Molecular Dynamics, Sunnyvale, CA) revealed that, on average, a 5-fold higher signal was observed in the glioblastoma multiforme tumors as compared with the anaplastic astrocytoma, and a 10-fold higher signal as compared with the mixed glioma, juvenile pilocytic astrocytomas, and dysembryoplastic neuroepithelial tumors. Results of TN immunohistochemistry in the astrocytomas correlated with glioma grade, with stronger staining of the hyperplastic vessels and tumor cells being observed in higher grade gliomas. No TN immunoreactivity was detected in the walls of the ependymomas, arachnoid cysts, and glial ependymal cyst that lack hyperplastic vessels, and minimal TN immunoreactivity was observed in the perivascular gliotic rim of the craniopharyngiomas. No TN was detected in the cyst fluid of these cystic processes. CONCLUSION: The presence of TN in and around the hyperplastic vessels and tumor cells present in the cyst walls of astrocytomas and its deposition in the intratumoral cyst fluid in which angiogenic factors have been detected further suggests a role for TN as an angiogenic modulator. These preliminary results suggest that immunodetection of TN in the tumor cyst fluid may indicate tumor type and grade.

Adolescent↗
PubMed1996

Migration of brain tumor cells on extracellular matrix proteins in vitro correlates with tumor type and grade and involves alphaV and beta1 integrins.

An important contributor to the malignancy of brain tumors is their ability to infiltrate the brain. Extracellular matrix molecules and cell adhesion molecules on cell surfaces play key roles in cell migration. In the present study, we used reaggregates of dissociated cells from freshly excised human brain tumors to analyze the migration of cells from human brain tumors of different types and grades on many different adhesion proteins adsorbed to glass substrates. Proteins were chosen based on their presence in normal or neoplastic nervous tissue, and included the extra-cellular matrix molecules fibronectin, collagens, fibrinogen, laminin, tenascin-C, thrombospondin, and the neuron-glia cell adhesion molecule, Ng-CAM. Cells from astrocytomas (n = 24) migrated on a variety of substrates, in contrast to cells from primitive neuroectodermal tumors cells (n=6), which only migrated well on laminin, fibronectin, or type IV collagen but not on the other substrates. Typically, migrating cells from astrocytomas of all grades had long, slender processes, were usually bipolar, and their cell bodies did not spread well on any substrate. Although there was variability in the migration of cells from astrocytomas of the same grade, cells from high-grade astrocytomas tended to migrate more extensively (42.3 +/- 4.7 micrometers/16 h: n = 16) than cells from lower grade astrocytomas (28.9 +/- 3.9 micrometers/16 h; P = 0.07; n = 8); the most striking differences were observed for collagen substrates, on which cells from lower grade astrocytomas migrated at very low levels (7.6 +/- 2 .6 micrometers/16 h) and cells from high-grade astrocytomas at higher levels (24.4 +/- 5.2 micrometers;P = 0.01). In contrast to primary cells from glioblastomas (n = 13), glioblastoma cell lines (n = 10) consistently spread on various substrates and migrated at high levels (69.5 +/- 7.6 versus 46.4 +/-5.7 micrometers/16 h; P = 0.03), in particular, on collagens (108.4 +/- 20.2 versus 28.0 +/- 6.1 micrometers/16 h; P= 0.001). Specific monoclonal antibodies to alphaV and beta1 integrin monomers completely inhibited the migration of astrocytoma cells on most substrates, suggesting that alphaV and beta1 integrins play a crucial role in brain tumor infiltration. These studies also suggest that although a large number of extracellular matrix molecules may promote tumor cell migration, disrupting the function of only a few tumor cell receptors may be critical for tumor infiltration in the brain.

Antibodies↗
PubMed1996

Expression of polypeptide variants of receptor-type protein tyrosine phosphatase beta: the secreted form, phosphacan, increases dramatically during embryonic development and modulates glial cell behavior in vitro.

Glial cells express three splicing variants of a receptor-type protein tyrosine phosphatase called RPTP beta. Two are receptor forms that differ in a large extracellular domain. The third is a secreted proteoglycan called phosphacan that lacks the cytoplasmic phosphatase domains. We have now identified, by immunoblotting, proteins corresponding to these three forms of RPTP beta in rat C6 glioma cells and brain. The short receptor form is much more prevalent than the full-length receptor in C6 glioma cells. Phosphacan is much more abundant than either of the receptor forms in rat brain, and its expression increases progressively during embryonic development, while the receptor forms show only moderate changes. In contrast to the long form and phosphacan that were detected as proteoglycans, the short receptor form, lacking the large alternatively spliced domain, was not detected as a chondroitin sulfate proteoglycan. We recently showed that phosphacan binds to the neuron-glia cell adhesion molecule, Ng-CAM, and we now report that glia expressing RPTP beta adhere and extend processes on substrates coated with Ng-CAM. After one day in culture, however, the glia retract their processes and often lift off the substrate. Conditioned medium from glial cells, which contains large amounts of phosphacan, inhibits glial adhesion to Ng-CAM, and depletion of phosphacan from the conditioned medium by immunoadsorption reduces the inhibitory activity. The results show that phosphacan increases dramatically during development, and indicate that secreted forms of RPTP beta can modulate glial cell adhesion and behavior.

Animals↗
PubMed1996

Tenascin-C expression by angiogenic vessels in human astrocytomas and by human brain endothelial cells in vitro.

The expression of the extracellular matrix glycoprotein tenascin-C (TN) is enhanced in human astrocytomas and correlates with angiogenesis. To determine whether vascular cells are able to synthesize TN, we investigated the expression of TN protein and mRNA in nine astrocytomas. Immunogold electron microscopy in two glioblastomas multiforme detected the presence of TN in an extracellular perivascular location and to a lesser extent among tumor cells, confirming light microscopy immunohistochemical findings. In situ hybridization of astrocytomas using a digoxigenin-labeled antisense riboprobe detected strong staining for TN mRNA in vascular cells, especially in hyperplastic vessels, including those at the invasive edge of the tumors but not in vessels of normal brains. We observed weaker staining in tumor cells indicating a higher level of TN mRNA in vascular than in tumor cells. No staining was detected with the sense probe. Moreover, we investigated the ability of human brain microvessel endothelial cells (HBMECs) in primary culture to synthesize TN in vitro. Western blot analysis of the culture supernatants from HBMECs detected large amounts of TN. Immunogold silver staining demonstrated the presence of TN on the surface of HBMECs and in the subendothelial matrix. The distribution of TN mRNA in vascular cells of astrocytomas and the ability of HBMECs to synthesize TN in vitro demonstrate that vascular cells, including endothelial cells, are a major source of TN associated with angiogenesis. Furthermore, our results suggest that TN expression may be associated with endothelial cell activation and may play an important role in angiogenesis.

Astrocytoma↗
PubMed1996

Functions of brain chondroitin sulfate proteoglycans during developments: interactions with adhesion molecules.

Chondroitin sulfate proteoglycans (CSPGs), including neurocan and phosphocan, are believed to be major components of brain extracellular matrix that interact with other matrix proteins and cell surface receptors. In addition, several brain CSPGs such as receptor protein tyrosine phosphatase beta are expressed as cell surface receptors that interact with proteins in the extracellular matrix and with receptors on neural cells. Recent in vitro studies demonstrate that, although the brain CSPGs neurocan and phosphocan can promote transient adhesion of neuronal cells, they inhibit stable cell adhesion and neurite growth promoted by the cell adhesion molecule Ng-CAM/L1. Neurocan and phosphocan bind with high affinity to Ng-CAM/L1 and N-CAM which may be their major receptors on neurons. These CSPGs also bind to other adhesion molecules, such as tenascin-C, and can differentially modulate adhesion of glia of tenascin-C. Both the glycosaminoglycan and the core glycoproteins contribute to the function of the brain CSPGs. When expressed in regions containing low levels of adhesion molecules, various CSPGs including phosphocan, neurocan, versican, aggrecan, and NG2 proteoglycan may act as barriers to cell migration and axonal growth. In regions containing high levels of adhesion proteins, brain CSPGs may still act to maintain certain boundaries while allowing selective axonal extension to proceed. There are numerous regions of overlap in the expression patterns of CSPGs and adhesion molecules in vivo, and the relative levels of these molecules as well as the organization of the extracellular matrix may be important factors that regulate the rate of axonal growth locally. Differential expression of CSPGs may be important for modulating cell adhesion as well as axonal growth and guidance during neural development, and continued expression may prevent these processes in the normal nature nervous system as well as following brain injury.

Aging↗
PubMed1995

The carbonic anhydrase domain of receptor tyrosine phosphatase beta is a functional ligand for the axonal cell recognition molecule contactin.

Receptor-type protein tyrosine phosphatase beta (RPTP beta) is expressed in the developing nervous system and contains a carbonic anhydrase (CAH) domain as well as a fibronectin type III repeat in its extracellular domain. Fusion proteins containing these domains were used to search for ligands of RPTP beta. The CAH domain bound specifically to a 140 kDa protein expressed on the surface of neuronal cells. Expression cloning in COS7 cells revealed that this protein is contactin, a GPI membrane-anchored neuronal cell recognition molecule. The CAH domain of RPTP beta induced cell adhesion and neurite growth of primary tectal neurons, and differentiation of neuroblastoma cells. These responses were blocked by antibodies against contactin, demonstrating that contactin is a neuronal receptor for RPTP beta. These experiments show that an individual domain of RPTP beta acts as a functional ligand for the neuronal receptor contactin. The interaction between contactin and RPTP beta may generate unidirectional or bidirectional signals during neural development.

Amino Acid Sequence↗
PubMed1995

Tenascin expression in astrocytomas correlates with angiogenesis.

We investigated the expression and distribution of the extracellular matrix protein tenascin (TN) in 59 astrocytomas and 11 samples of normal brain by Western blot analysis and immunohistochemistry using antibodies against human TN. The tumors included 14 juvenile pilocytic astrocytomas (grade 1), 13 low grade fibrillary astrocytomas (grade II), 8 anaplastic astrocytomas (grade III), and 24 glioblastomas multiforme (grade IV). Proliferation indices were calculated by computer-based image analysis after immunostaining with the MIB-1 antibody against the Ki-67 proliferation-associated antigen. Western blot analysis for TN on fresh frozen tumor tissue from 23 of the 59 astrocytomas indicated up to 4-fold higher TN expression in glioblastomas multiforme than in nontumorous control tissues. Enhanced intercellular expression of TN was observed by immunohistochemistry in glioblastomas multiforme. More-over, TN immunostaining was consistently greater within and around the walls of hyperplastic blood vessels than nonhyperplastic vessels of both high grade tumors and juvenile pilocytic astrocytomas. Juvenile pilocytic astrocytomas with increased TN expression by Western blot analysis had vascular hyperplasia by light microscopy. Proliferation indices moderately correlated with tumor grade. Enhanced immunohistochemical expression of TN was associated with higher tumor grade with higher proliferation indices. The strong association of TN and vascular hyperplasia, regardless of tumor grade, suggests that TN may play a crucial role in angiogenesis.

Astrocytoma↗
PubMed1994

Biochemical characterization and immunolocalization of SC2 protein: SC2 protein is indistinguishable from the cell adhesion molecule axonin-1.

SC2 is a monoclonal antibody that was previously shown to recognize a subset of neurons in the developing nervous system of the chick. We have now used the SC2 monoclonal antibody to purify from chick embryo brain membranes a glycoprotein that migrates at approximately 125 kDa on SDS/PAGE. The size of this protein and its distribution pattern in the spinal cord are similar to that observed for axonin-1. A polyclonal anti-axonin-1 antibody R26 specifically reacted with the SC2 protein from brain. This antibody, as well as polyclonal antibody (369) against purified SC2 protein, reacted with 115-130 kDa proteins in vitreous humor, a rich source of axonin-1, and with similar sized proteins precipitated from vitreous humor by the 369, and SC2 antibodies. Treatment of SC2 protein isolated from chick brain membranes with PI-PLC indicated that it contains a glycophosphatidylinositol (GPI) moiety. Co-aggregation experiments using Covaspheres with covalently bound proteins indicated that SC2 protein binds heterophilically to Ng-CAM. Immunohistochemical analysis of chick embryos showed that SC2 protein is abundant in the sensory nerve bundles of both the central and peripheral nervous systems during development. Its expression was restricted and it was specifically localized in the dorsal funiculus of the spinal cord, as well as in olfactory, retinal, trigeminal, vestibulocochlear, glossopharyngeal and vagal nerve fibers. The biochemical and immunohistochemical data show that SC2 protein is axonin-1, and the immunolocalization studies support the hypothesis that SC2 protein may play a role during development of particular fiber systems by interacting with other cell adhesion molecules such as Ng-CAM.

Animals↗
PubMed1994

Receptor tyrosine phosphatase beta is expressed in the form of proteoglycan and binds to the extracellular matrix protein tenascin.

The extracellular domain of receptor type protein tyrosine phosphatase beta (RPTP beta) exhibits striking sequence similarity with a soluble, rat brain chondroitin sulfate proteoglycan (3F8 PG). Immunoprecipitation experiments of cells transfected with RPTP beta expression vector and metabolically labeled with [35S]sulfate and [35S]methionine indicate that the transmembrane form of RPTP beta is indeed a chondroitin sulfate proteoglycan. The 3F8 PG is therefore a variant form composed of the entire extracellular domain of RPTP beta probably generated by alternative RNA splicing. Previous immunohistochemical studies indicated that both RPTP beta and the extracellular matrix protein tenascin are localized in similar regions of the central nervous system. We have performed co-aggregation assays with red and green Co-vaspheres coated with tenascin and 3F8 PG, respectively, showing that the extracellular domain of RPTP beta (3F8 PG) binds specifically to tenascin. The interaction between a receptor tyrosine phosphatase and an extracellular matrix protein may have a role in development of the mammalian central nervous system.

Alternative Splicing↗
PubMed1994

Interactions with tenascin and differential effects on cell adhesion of neurocan and phosphacan, two major chondroitin sulfate proteoglycans of nervous tissue.

We have studied interactions of tenascin with two chondroitin sulfate proteoglycans, neurocan and phosphacan. Neurocan is a multi-domain proteoglycan with a 136-kDa core protein that is synthesized by neurons and binds to hyaluronic acid, whereas the 173-kDa core protein of phosphacan, which is synthesized by glia, represents an extracellular variant of the receptor-type protein tyrosine phosphatase RPTP zeta/beta. Keratan sulfate-containing glycoforms of phosphacan (designated phosphacan-KS) are also present in brain. Immunocytochemical studies of early postnatal rat cerebellum demonstrated that the localization of neurocan, phosphacan, and phosphacan-KS all overlap extensively with that of tenascin, an extracellular matrix protein that modulates cell adhesion and migration. Binding studies using purified proteins covalently attached to fluorescent microbeads demonstrated that proteoglycan-coated beads co-aggregated with differently fluorescing beads coated with tenascin. The co-aggregation was specifically inhibited by Fab' fragments of antibodies against tenascin or the proteoglycans and by soluble neurocan, phosphacan, and tenascin. A solid phase radioligand binding assay confirmed that neurocan, phosphacan, and phosphacan-KS bind to tenascin but not to laminin and fibronectin. Chondroitinase treatment of the proteoglycans or addition of free chondroitin sulfate had no significant effect, indicating that the binding activity is mediated largely via the core glycoproteins. Scatchard analysis demonstrated high affinity binding of 125I-phosphacan, phosphacan-KS, and neurocan to a single site in tenascin, and neurocan and various glycoforms of phosphacan all inhibited binding of 125I-phosphacan to tenascin. In studies of cell adhesion to proteins adsorbed to Petri dishes, phosphacan inhibited adhesion of C6 glioma cells to tenascin whereas neurocan had no effect. Our results suggest that tenascin binds phosphacan and neurocan in vivo and that interactions between chondroitin sulfate proteoglycans and tenascin may play important roles in nervous tissue histogenesis, possibly by modulating signal transduction across the plasma membrane.

Animals↗
PubMed1994

Regions of the human neurokinin A receptor involved in the generation of second messengers and in receptor desensitization.

Deletion analysis was used to study sites of human Neurokinin A receptor (HNKAR) necessary for signal transduction in CHO cells. Deletion of 62 and 81 amino acids from the c-terminus of HNKAR forms mutant receptors HNKAR delta 62 and HNKAR delta 81, which bind neurokinin A with high affinity but are functionally different. Wild type HNKAR and HNKAR delta 62 are functionally active whereas HNKAR delta 81 is functionally inactive. In addition, HNKAR and HNKAR delta 62 were both desensitized to the neurokinin A signal within 5 minutes. The data indicates: 1) an intact cytoplasmic tail of the HNKAR is not critical for signal transduction, but the n-terminal amino acids of the cytoplasmic tail are necessary for signaling and 2) the c-terminal 62 amino acids are not necessary for desensitization.

Amino Acid Sequence↗