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C Arregui

Publications and source records attributed to C Arregui.

11 recordsLinked to original sources
PubMed2001

PTP1B regulates neurite extension mediated by cell-cell and cell-matrix adhesion molecules.

N-cadherin and beta1-integrin adhesion and signaling play important roles in growth cone adhesion and guidance. Each of these adhesion receptor systems is composed of multiprotein complexes, and both adhesion and downstream signaling events are regulated through the interaction of protein tyrosine kinases and phosphatases with many of the proteins that make up these complex systems. Work from our laboratory reported that the nonreceptor protein tyrosine phosphatase PTP1B is localized to adherens junctions and focal adhesion complexes and regulates both N-cadherin- and beta1-integrin-mediated adhesion. PTP1B appears to modulate integrin-mediated adhesion through regulation of src activation and cadherin-mediated adhesion through dephosphorylation of beta-catenin. We have continued these studies and report that PTP1B is localized to the tips of growing neurites and that introduction of a noncatalytic mutant of PTP1B into PC12 cells results in inhibition of N-cadherin- and beta1-integrin-mediated neurite outgrowth but is without effect on neurite outgrowth on poly-L-lysine. Moreover, suppressing the level of PTP1B in primary embryonic chick neural retina cells using antisense oligonucleotides also inhibits N-cadherin- and beta1-integrin-mediated neurite outgrowth. Neither of these techniques reduces the levels of expression of either adhesion receptor. We conclude that PTP1B is a regulatory component of the molecular complex required for both N-cadherin and beta1-integrin-mediated axon growth.

Animals↗
PubMed2000

The nonreceptor tyrosine kinase fer mediates cross-talk between N-cadherin and beta1-integrins.

Cadherins and integrins must function in a coordinated manner to effectively mediate the cellular interactions essential for development. We hypothesized that exchange of proteins associated with their cytoplasmic domains may play a role in coordinating function. To test this idea, we used Trojan peptides to introduce into cells and tissues peptide sequences designed to compete for the interaction of specific effectors with the cytoplasmic domain of N-cadherin, and assayed their effect on cadherin- and integrin-mediated adhesion and neurite outgrowth. We show that a peptide mimicking the juxtamembrane (JMP) region of the cytoplasmic domain of N-cadherin results in inhibition of N-cadherin and beta1-integrin function. The effect of JMP on beta1-integrin function depends on the expression of N-cadherin and is independent of transcription or translation. Treatment of cells with JMP results in the release of the nonreceptor tyrosine kinase Fer from the cadherin complex and its accumulation in the integrin complex. A peptide that mimics the first coiled-coil domain of Fer prevents Fer accumulation in the integrin complex and reverses the inhibitory effect of JMP. These findings suggest a new mechanism through which N-cadherin and beta1-integrins are coordinately regulated: loss of an effector from the cytoplasmic domain of N-cadherin and gain of that effector by the beta1-integrin complex.

Amino Acid Sequence↗
PubMed1999

The juxtamembrane domain of cadherin regulates integrin-mediated adhesion and neurite outgrowth.

Axons are guided along their trajectories during development by many different systems of adhesion, attraction, and repulsion. Thus, many distinct, and potentially competing, receptor systems respond to environmental cues, and the information must be coordinated inside the growth cone to ensure that extension follows the appropriate path. In this brief review we bring together two studies, each of which has defined different aspects of a pathway through which N-cadherin regulates beta1-integrin function allowing for coordinated responses to environmental cues during neurite extension. First we review progress in defining the binding to cells and the subsequent effects on adhesion and neurite outgrowth of the chondroitin sulfate proteoglycan, neurocan. Neurocan binds to a cell surface glycosyltransferase associated with N-cadherin (but not integrin), initiating a signal which results in loss of cadherin and integrin-function-suggesting that these two adhesion receptor systems engage in cross-talk, allowing coordinate regulation. Second, we review the use of "Trojan" peptides, peptides which mimic specific sequences in the cytoplasmic domain of N-cadherin attached to a cell permeation sequence, to reveal protein-protein interactions critical to cadherin-integrin cross-talk. One peptide mimicking a 20 amino acid sequence in the juxtamembrane region of N-cadherin has the same effect as neurocan, blocking both cadherin- and integrin-mediated adhesion and neurite outgrowth. Both neurocan and the peptide cause the release of the non-receptor tyrosine kinase Fer from the cadherin complex and its binding to the integrin complex. These data define an epigenetic pathway through which environmental cues are capable of coordinately regulating the activity of two developmentally important adhesion systems.

Animals↗
PubMed1999

Modulation of urokinase-type plasminogen activator and metalloproteinase activities in cultured mouse mammary-carcinoma cells: enhancement by paclitaxel and inhibition by nocodazole.

Paclitaxel is a potent anti-tumor drug used in the treatment of breast cancer. It induces de-centralization of the microtubular system in tumor cells, blocking cell division. In the search for dissemination to a secondary site, cancer cells are capable of degrading most components of the extracellular matrix via an extracellular proteolytic cascade, including urokinase-type plasminogen activator (uPA) and the matrix metalloproteinases (MMPs). In the present study, the effects of paclitaxel and nocodazole, 2 drugs known to affect microtubules with opposite mechanisms of action, have been tested for their effect on the secretion of uPA and MMPs in cultures of F3II mouse mammary-tumor cells. Tumor-derived uPA activity significantly increased after pre-treatment of tumor cells for 24 hr with micromolar concentrations of paclitaxel (4 microM), while decreasing after pre-treatment with nocodazole (1 microM). A similar modulation was found for MMP-9 by zymographic analysis. Immunofluorescence and Western-blot analysis confirmed the formation of parallel microtubule fragments in paclitaxel-treated cells and almost complete de-polymerization of microtubules in nocodazole-treated ones. Our data suggest that, through opposite actions on microtubule organization and dynamics, paclitaxel and nocodazole exert inverse modulation of tumor-derived proteolytic activity in mammary tumor cells.

Animals↗
PubMed1998

The nonreceptor protein tyrosine phosphatase PTP1B binds to the cytoplasmic domain of N-cadherin and regulates the cadherin-actin linkage.

Cadherin-mediated adhesion depends on the association of its cytoplasmic domain with the actin-containing cytoskeleton. This interaction is mediated by a group of cytoplasmic proteins: alpha-and beta- or gamma- catenin. Phosphorylation of beta-catenin on tyrosine residues plays a role in controlling this association and, therefore, cadherin function. Previous work from our laboratory suggested that a nonreceptor protein tyrosine phosphatase, bound to the cytoplasmic domain of N-cadherin, is responsible for removing tyrosine-bound phosphate residues from beta-catenin, thus maintaining the cadherin-actin connection (). Here we report the molecular cloning of the cadherin-associated tyrosine phosphatase and identify it as PTP1B. To definitively establish a causal relationship between the function of cadherin-bound PTP1B and cadherin-mediated adhesion, we tested the effect of expressing a catalytically inactive form of PTP1B in L cells constitutively expressing N-cadherin. We find that expression of the catalytically inactive PTP1B results in reduced cadherin-mediated adhesion. Furthermore, cadherin is uncoupled from its association with actin, and beta-catenin shows increased phosphorylation on tyrosine residues when compared with parental cells or cells transfected with the wild-type PTP1B. Both the transfected wild-type and the mutant PTP1B are found associated with N-cadherin, and recombinant mutant PTP1B binds to N-cadherin in vitro, indicating that the catalytically inactive form acts as a dominant negative, displacing endogenous PTP1B, and rendering cadherin nonfunctional. Our results demonstrate a role for PTP1B in regulating cadherin-mediated cell adhesion.

Actins↗
PubMed1997

A phospholipase D and protein kinase C inhibitor blocks the spreading of murine mammary adenocarcinoma cells altering f-actin and beta1-integrin point contact distribution.

Spreading is a critical process involved in motility and growth of tumor cells during the metastatic cascade. Focal adhesion kinase, src-proteins and PKC have been reported to participate in the regulation of cytoskeleton organization in both normal and transformed cells during spreading. The role of other signaling enzymes such as PLD and PAP has not been studied during spreading in tumor cells. We now show that the spreading of murine mammary adenocarcinoma LM3 cells was significantly reduced by n-butanol, a PLD and PKC inhibitor, with a maximal inhibition of 54% (p < 0.001) in both the presence and absence of serum, as measured by phase-contrast microscopy. PMA only stimulated cell spreading over the control in the absence of serum and n-butanol inhibition was completely reversed by PMA treatment in both conditions. PA, the product of PLD activity, stimulated LM3 cell spreading and the same effect was observed with staurosporine. Spreading was enhanced when cells were seeded on collagen-IV- or fibronectin-coated surfaces and n-butanol could inhibit both integrin-derived signals. Cell spreading inhibition correlated with the absence of f-actin bundles and fewer beta1-integrin point contacts as determined by double immunofluorescence microscopy. In addition, n-butanol inhibited the proliferation of LM3 cells in the presence of serum (p < 0.01). These results suggest that beta1-integrin and f-actin/point contact assembly, involved in spreading and proliferation, require the participation of PLD-PKC regulatory pathways in LM3 cells.

1-Butanol↗
PubMed1992

Peanut agglutinin binding glycoproteins in the chick retina: their presence in Müller glia cells.

The histological and cellular distribution and some biochemical characteristics of components that bind peanut agglutinin (PNA), a lectin that recognizes preferentially terminal galactose-beta (1-3) N-acetyl galactosamine disaccharide residues of glycoconjugates, were studied in chick retinal tissue and in dissociated retinal cells after their differentiation in culture. In sections of retinal tissue from animals 7 days after hatching (Rp7), in addition to the inner and outer segments of the photoreceptor layer, the plexiform and optic fiber layers were stained with rhodamine-labeled PNA, indicating that, besides photoreceptor cells, other cellular types contribute to the PNA staining. We present evidence indicating that at least part of this staining is provided by Müller glia cells. In cultures of dissociated cells from retinas at embryonic day 7 (R7), photoreceptor-like cells and flat Müller glia-derived cells but not neurons were stained with rhodamine-labeled PNA. Furthermore, Müller glia cells isolated from Rp7 were also brightly stained with PNA. Western blot assays of extracts from R7 showed the presence of PNA binding glycoproteins of 31-33 kDa and a component that migrates at the dye front. In addition to the components detected in R7, extracts from R14 and Rp7 showed the presence of a major PNA binding glycoprotein of 175 kDa and a minor glycoprotein of 220 kDa. Extracts from the photoreceptor layer contain the 175 and 220 kDa glycoproteins, indicating their association with photoreceptor cells. The 31-33 kDa components were detected in extracts from the remnant inner retina, suggesting their association with the Müller glia cells. Supporting this view, these components and not those of 175 and 220 kDa were detected in cell cultures enriched in flat Müller glia-derived cells. Only the 31-33 kDa components and the component that migrates at the dye front were detected in extracts from cell cultures enriched in photoreceptor-like cells, suggesting the need of some environmental element for the expression of the 175 and 220 kDa components in the differentiated photoreceptor cells.

Animals↗
PubMed1991

Tyrosinated and detyrosinated microtubules in axonal processes of cerebellar macroneurons grown in culture.

We have used the monoclonal antibody YL 1/2 (Tyr) specific for tyrosinated tubulin, and a polyclonal antibody (Glu) specific for detyrosinated tubulin to visualize the distribution of microtubules and microtubule assembly sites during axonal outgrowth. Cerebellar macroneurons growing in culture initially extend several short and thin neurites which have the potential to differentiate either as axons or dendrites (Ferreira and Caceres: Developmental Brain Research 49:205-213, 1989). At the onset of axonal outgrowth the Tyr antibody labels the minor neurites, the axon, and its growth cone, while the Glu antibody only shows immunoreactivity in the axonal shaft. After nocodazole treatment, the Tyr staining disappears, whereas that produced by the Glu antibody remains practically unchanged. When nocodazole was removed, tyrosinated microtubules reappeared first at the tip of the axon, in a more distal region than that occupied by detyrosinated microtubules; another focal site of tyrosinated tubulin incorporation was detected in the cell body. Incorporation of tyrosinated tubulin into growing axons was also studied after taxol treatment. After long incubation periods in the presence of taxol, the Tyr staining disappeared from the axon but remained in the cell body; however, immunoreactivity in this site was negative when the cells were preincubated in the presence of protein synthesis inhibitors. Release from taxol results in the reappearance of Tyr immunoreactivity at the distal end of the axon. Taken collectively, the present results indicate 1) that in cerebellar macroneurons axonal differentiation is accompanied by a temporal and spatial differentiation of microtubules and 2) that there is an active site of tyrosinated tubulin assembly at the tip of axonal processes, and they suggest that the highly tyrosinated domain in this region is a consequence of rapid microtubule turnover and tubulin tyrosine ligase activity.

Alkaloids↗
PubMed1990

Specific antibodies for tyrosinated and detyrosinated tubulin recognize retina tubulin subpopulations that do not participate in the posttranslational tyrosination/detyrosination cycle.

We have used the monoclonal YL 1/2 (Tyr antibody) and polyclonal (Glu antibody) antibodies, specific for tyrosinated and detyrosinated tubulin, respectively, to determine the levels and cellular distribution of these tubulin species in chick retina during development. At embryonic day 4, detyrosinated tubulin was restricted to the ganglion cells of the fundic region. As development progresses, immunofluorescence also appears, first, in the outermost zone of the retina and then in the plexiform layers. The Tyr antibody staining was found in the different layers and it was fairly homogeneous in distribution. Analysis by dot immunobinding showed that the ratios of tyrosinated to detyrosinated tubulin obtained at different ages do not agree with those obtained previously by an enzymatic method based on the incorporation of [14C]tyrosine. We found that the lack of coincidence is due to the fact that a fraction of the tubulin species determined by the Tyr and Glu antibodies does not participate in the posttranslational tyrosination/detyrosination cycle. This is a novel concept that should be considered in the interpretations of immunofluorescence studies concerning the cellular distribution of tyrosinated and detyrosinated tubulin.

Animals↗
PubMed1989

Tyrosination state of tubulin and the activity of tubulin:tyrosine ligase and tubulin carboxypeptidase in the developing retina of the chick.

The tyrosination state of tubulin and the enzymes involved in the tubulin tyrosination/detyrosination cycle--tubulin:tyrosine ligase and tubulin carboxypeptidase--were determined in chick retina during development. The amount of tyrosinable (tyrosinated plus detyrosinated) tubulin increased approximately 110% from embryonic day 7 to 14. Then it decreased, and by day 19 it was similar to the value on day 7. This result did not change after hatching, at least up to day 20. The proportion of tyrosinated and detyrosinated tubulin significantly changed with the development of the animal. At embryonic day 7, these tubulin species were at a proportion of 70 and 30%, respectively, and after hatching, the values inverted, to 30 and 70%, respectively. This change did not correlate with the activity of the ligase relative to that of the carboxypeptidase, as measured in vitro. This observation suggested that a change in the turnover rate of microtubules, in the proportion of assembled and nonassembled tubulin pools, or in both had occurred. Coincident with the last possibility, the proportion of assembled tubulin was found to increase during the development of the animal. This finding suggests that the tyrosination state of tubulin may be determined, at least in part, by the assembly state.

Animals↗