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K Meiri

Publications and source records attributed to K Meiri.

7 recordsLinked to original sources
PubMed1997

Cell signalling and CAM-mediated neurite outgrowth.

A wide range of molecules promote nerve growth, and these include cell adhesion molecules (CAMs), NCAM, N-cadherin, and the L1 glycoprotein are CAMs that are normally found on both the advancing growth cone and also on cellular substrates, and in general operate via a homophilic binding mechanism. In recent years it has become clear that nerve growth stimulated by these CAMs does not rely on the adhesion function of these molecules, but instead requires that the CAMs activate second messenger cascades in neurons. A large body of evidence supports the hypothesis that homophilic binding of the CAM in the substrate to the CAM in the neuron leads to activation of the neuronal FGF receptor, possibly via a direct interaction in cis between the CAM and the FGF receptor. The consequential activation of PLC gamma is both necessary and sufficient to account for the neurite outgrowth response stimulated by the above three CAMs. Based on the above model, we reasoned that soluble CAMs might also be able to stimulate neurite outgrowth and that such agents might be developed as potential therapeutic agents for stimulating nerve regeneration. To this end we have made soluble chimeric molecules consisting of the extracellular domain of NCAM or L1 fused to the Fc region of human IgG 1. We have found that these molecules can stimulate neurite outgrowth from rat and mouse cerebellar granule cells cultured on a variety of tissue culture substrates and that they do so by activating the FGF receptor signal transduction cascade in the neurons. Consistent with this model, we find that neurons that have their FGF receptor function ablated as a consequence of the expression of dominant negative FGF receptors, no longer respond to the soluble CAM. Downstream targets of CAM function have also been studied. Addition of soluble CAMs to isolated growth cone preparations from mouse or rat brain leads to enhanced phosphorylation of the GAP-43 protein providing a link between the cell surface and the cytoskeleton.

Animals↗
PubMed1995

The earliest patterns of neuronal differentiation and migration in the mammalian central nervous system.

With the use of four independent cell markers and Brd-U birthdating we have charted the earliest stages of neuronal differentiation and migration in the developing rat central nervous system, including the cortex, spinal cord, and retina. One of the markers, the monoclonal antibody 2G12, labeled a large subpopulation of differentiating cells that uniformly lined the ventricles throughout these CNS regions at unexpectedly early ages. Immunocytochemistry demonstrated that, in cortex, the 2G12 antigen could appear in cells during mitosis. More mature looking 2G12-positive cell types initially had a primitive radial morphology and were axonless. However, in strict spatio-temporal sequences, the most mature looking 2G12-positive cells had the ability to sprout GAP-43-positive axons before or after the cell body left the ventricular surface and before or after detachment of their pial or ventricular endfoot processes. Double label experiments with 2G12 and Brd-U showed that none of these three 2G12-positive cell types incorporated Brd-U after a short pulse. The primitive neuroepithelial shape of the immature neurons was verified with a polyclonal GAP-43 antibody, a type III beta tubulin antibody, and DiI labeling from a distal portion of the axon. In the cortex and retina, the 2G12 marker persisted in cells that had reached prospective neuronal layers. However, in all CNS regions observed, 2G12 immunoreactivity disappeared from the cell body as the axon extended from the young neuron. Based on the smooth progression of changing 2G12-positive cell shapes, but also because of the transient nature of this label, we can only speculate that the 2G12 epitope may be marking a continuum of neuronal cell states throughout the earliest period of differentiation and migration. Thus, our hypothesis suggests that many of the youngest CNS neurons may have a widespread distribution and may begin their differentiation, and even remain axonless for a time, while retaining a neuroepithelial morphology. Once differentiation resumes, a major mode of transformation into mature neurons during the earliest stages of development could occur via translocation of the cell soma into the pial process. Importantly, these markers have verified at later stages, and especially in cortex, that multiple mechanisms exist for neuronal migration in the CNS depending on the region and stage of development.

Animals↗
PubMed1993

Developmental changes of growth cone gangliosides of the postnatal rat cerebrum.

The ganglioside content and composition of growth cones prepared from forebrains of day 0 (p0), day 2 (p2), and day 6 (p6) postnatal rats was analysed in comparison with that of whole forebrain membranes obtained from rats of the same age. The amount of total ganglioside bound sialic acid (microgram NeuAc mg protein-1) was found to increase in both whole cerebrum membranes and growth cones from p0 to p6. The ganglioside concentrations of growth cones exceeded that of the whole cerebrum by 35-40% at all time points investigated. The major rat brain gangliosides were also found in the growth cone fractions, however, in different relative amounts. Thus, the well known developmental shift from the b- to the a-ganglioside synthesis pathway was significantly delayed in growth cones, indicating that this shift is not associated with axonal growth but more likely with the subsequent period of synaptogenesis. GQ1c, recognized by the mAb Q211, and an unidentified growth-cone specific ganglioside x were found to be expressed in growth cones of p0 rats and to decrease up to p6.

Animals↗
PubMed1992

Purification and lectin-binding properties of s-laminin, a synaptic isoform of the laminin B1 chain.

The extracellular matrix (ECM) at the vertebrate neuromuscular junction is a repository of functionally important molecules, some of which can regulate the formation of synapses during regeneration. One candidate molecule is s-laminin, a 185-kDa homologue of the laminin B1 chain. Whereas several members of the laminin family are present throughout the ECM ensheathing muscle fibers, immunoreactivity for s-laminin is found selectively at synaptic sites in adult and embryonic rats, and is detectable at a time when synaptogenesis is taking place during development. We have reported previously that a rat schwannoma cell line, D6P2T, produces and releases large amounts of s-laminin in culture. We have now purified s-laminin from medium conditioned by these cells by using a simple three-step procedure. Serum-free, conditioned medium is separated by ion-exchange chromatography on DEAE-Sephacel, followed by size-exclusion chromatography on 500 HR-Sephacryl. Finally, s-laminin is dissociated from other ECM components by agarose gel electrophoresis under reducing conditions and recovered in solution by extracting slices of agarose gel. The purified preparation displays one silver-stained band that is recognized by three monoclonal antibodies known to bind to different epitopes on s-laminin. Lectin-binding studies demonstrate that s-laminin is a glycoprotein and bears many of the carbohydrate moieties present on the B1 and B2 chains of laminin. Thus, the three 185-220-kDa members of the laminin family are related in both their protein and carbohydrate domains.

Animals↗
PubMed1990

Purification of the growth-associated protein GAP-43 by reversed phase chromatography: amino acid sequence analysis and cDNA identification.

GAP-43 is a neuronal phosphoprotein. Increased synthesis and axonal transport of GAP-43 has been associated with axon growth, and altered phosphorylation of GAP-43 has been associated with changes in synaptic efficacy. Here we report a rapid and effective procedure employing reverse-phase HPLC for the purification of GAP-43 from rat brain. To characterize the protein purified by this procedure, we generated proteolytic fragments and determined their amino acid sequences. These directly determined sequences, corresponding to 56% of the GAP-43 amino acids, confirm recently reported sequences deduced from the nucleotide sequences of cDNAs. Using oligonucleotide probes constructed according to these amino acid sequences, we identified GAP-43 cDNAs in a library prepared from neonatal rat superior cervical ganglion cells. One of these cDNAs was 1.1 kB in size; it hybridized specifically with a 1.5 kB RNA from brain, but not from liver, and contained the entire coding sequence for GAP-43. This cDNA differed from recently reported cDNAs in its 3' untranslated region.

Amino Acid Sequence↗
PubMed1989

A GAP-43-like protein in cat visual cortex.

We have purified a protein that changes in relative concentration during the development of the kitten visual cortex. It resembles GAP-43 (a neuronal protein that is expressed at elevated levels during periods of development and regenerative axon growth) in the following respects: (1) it is an acidic protein (pI = 4.7) whose electrophoretic mobility on SDS-PAGE is similar to, but lower than rat GAP-43, suggesting that the cat protein is larger; (2) its electrophoretic mobility varies with the acrylamide concentration in a manner that is characteristic of GAP-43; (3) its concentration in kitten forebrain is elevated during early postnatal development; (4) the sequence of ten consecutive amino acids from a chemically generated fragment matches the expected sequence from GAP-43; and (5) its amino-acid content also matches GAP-43. We conclude that our purified protein is cat GAP-43. Immunoblots with an antibody prepared against rat GAP-43 suggested that the concentration of GAP-43 in the visual cortex declines with age.

Acrylamide↗