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M C Fishman

Publications and source records attributed to M C Fishman.

At least 127 records · Page 7Linked to original sources
PubMed1992

Patterning the zebrafish heart tube: acquisition of anteroposterior polarity.

The patterning of an internal organ, like the heart, is little understood. Central to this patterning is the formation, or the acquisition, of an anteroposterior (A-P) axis. We have approached the question of how the heart tube acquires polarity in the zebrafish, Brachydanio rerio, which offers numerous advantages for studying cardiac morphogenesis. During the early stages of organogenesis in the fish, the heart tube lies in an A-P orientation with the venous end lying anteriorly and the arterial end lying posteriorly. High doses (10(-6)-10(-5)M) of retinoic acid (RA) cause truncation of the body axis, as they do in Xenopus. Low doses of retinoic acid (10(-8)-10(-7) M), which do not appear to affect the rest of the embryo, have pronounced effects upon heart tube morphogenesis, causing it to shrink progressively along the A-P axis. To investigate this further, we identified monoclonal antibodies that distinguish between the zebrafish cardiac chambers and used them to show that the RA-induced cardiac truncation always begins at the arterial end of the heart tube. There is a continuous gradient of sensitivity from the arterial to the venous end, such that increasing RA exposure causes the progressive and sequential deletion first of the bulbus arteriosus and then, in order, of the ventricle, the atrium, and the sinus venosus. As exposure increases, parts of chambers are deleted before entire chambers; thus, the sensitivity to RA appears to be independent of chamber boundaries. The analysis of the heart tube's sensitivity to RA and its timing suggest that polarity is established during or shortly after initial commitment to the cardiac lineage.

Animals↗
PubMed1991

An intracellular guanine nucleotide release protein for G0. GAP-43 stimulates isolated alpha subunits by a novel mechanism.

G protein-coupled membrane receptors activate G proteins by enhancing guanine nucleotide exchange. G0 is a major component of the growing regions (growth cones) of neurons. GAP-43 is a neuronal protein associated with the cytosolic face of the growth cone plasma membrane and stimulates binding of guanosine 5'-3-O-(thio)triphosphate (GTP gamma S) to Go (Strittmatter, S. M., Valenzuela, D., Kennedy, T. E., Neer, E. J., and Fishman, M. C. (1990) Nature 344, 836-841). Here we have examined the mechanism by which GAP-43 affects G0. Like G protein-coupled receptors, GAP-43 enhances GDP release from G0, increases the initial rate of GTP gamma S binding, and increases the GTPase activity of Go, all without altering the intrinsic kappa cat for the GTPase. Unlike the case for receptors, however, the GAP-43 effect is not blocked by pertussis toxin, nor affected by the presence or absence of beta gamma or of phospholipids. There is specificity to the interaction, in that GAP-43 increases GTP gamma S binding to recombinant alpha o and alpha i1, but not to recombinant alpha s. Thus, GAP-43 is a guanine nucleotide release protein with a novel mechanism of action, potentially controlling membrane-associated G proteins from within the cell.

Animals↗
PubMed1991

The neuronal growth cone as a specialized transduction system.

Neuronal growth and remodelling are guided by both intracellular gene programs and extracellular stimuli. The growth cone is one site where the effects of these extrinsic and intrinsic factors converge upon the mechanical determinants of cell shape. We review the growth cone as a transduction device, converting extracellular signals into mechanical forces. A variety of soluble, extracellular matrix and membrane bound molecules control growth cone behavior. In addition, GAP-43 is discussed as a possible component of the intraneuronal gene program which modulates growth cone activity. The GTP-binding protein, Go, is a major growth cone membrane protein that may transduce signals not only from outside the cell, but from within as well. This may provide a molecular site in the growth cone for the coordination of a genetic growth program with environmental signals.

Animals↗
PubMed1991

Growth cone transduction: Go and GAP-43.

The neuronal growth cone plays a crucial role in forming the complex brain architecture achieved during development, and similar nerve terminal mechanisms may operate to modify synaptic structure during adulthood. The growth cone leads the elongating axon towards appropriate synaptic targets by altering motility in response to a variety of extracellular signals. Independently of extrinsic clues, neurons manifest intrinsic control of their growth and form (Banker and Cowan, 1979). Hence, there must be intracellular proteins which control nerve cell shape, so-called 'plasticity' or 'growth' genes. GAP-43 may be such a molecule (Skene and Willard, 1981; Benowitz and Lewis, 1983). For example, GAP-43 is localized to the growth cone membrane (Meiri et al. 1986; Skene et al. 1986) and can enhance filopodial formation even in non-neuronal cells (Zuber et al. 1989a). It includes a small region at the amino terminus for membrane association and perhaps growth cone targeting (Zuber et al. 1989b, Liu et al. 1991). We have found that Go, a member of the G protein family that links receptors and second messengers, is the major non-cytoskeletal protein in the growth cone membrane (Strittmatter et al. 1990). Double staining immunohistochemistry for GAP-43 and Go shows that the distributions of the two proteins are quite similar. Purified GAP-43 regulates the activity of purified Go (Strittmatter et al. 1990), a surprising observation since GAP-43 is an intracellular protein. We have compared the mechanism of GAP-43 activation of Go with that of G protein-linked receptors.2+ interactions between Go and GAP-43 suggest that Go plays a pivotal role in growth cone function, coordinating the effects of both extracellular signals and intracellular growth proteins.

Animals↗
PubMed1991

SV40 T antigen transforms calcitonin cells of the thyroid but not CGRP-containing neurons in transgenic mice.

Neurons are postmitotic for the adult life of animals. Tumors rarely, if ever, arise from neurons in the adult, although they do from other cells from the same lineage, such as the neuroendocrine C cells of the thyroid. We have found that 2 kb of the calcitonin gene-related peptide (CGRP)/calcitonin gene suffices to target expression to CGRP-containing neurons, such as those in the dorsal root ganglia (DRG), and to the calcitonin-secreting C cells of the thyroid. Using this promoter we have examined the effect of two potentially transforming oncogenes in these two different populations. Overexpression of c-myc for periods of up to two years does not transform either cell type, whereas SV40 Tag causes early onset medullary thyroid carcinoma, but does not transform the dorsal root ganglia neurons. This suggests that as part of the terminal differentiation process of these neurons, the cessation of mitosis is accompanied by a relative refractoriness to oncogenes that may transform other cells of the same lineage.

Animals↗
PubMed1990

G0 is a major growth cone protein subject to regulation by GAP-43.

G0, a GTP-binding protein that transduces information from transmembrane receptors, has been found to be a major component of the neuronal growth cone membrane. GAP-43, an intracellular growth cone protein closely associated with neuronal growth, stimulates GTP-gamma-S binding to G0. It does so through an amino-terminal domain homologous to G-linked transmembrane receptors. Thus, G0 in the growth cone may be regulated by intracellular as well as extracellular signals.

Amino Acid Sequence↗
PubMed1989

A membrane-targeting signal in the amino terminus of the neuronal protein GAP-43.

Neurons and other cells, such as those of epithelia, accumulate particular proteins in spatially discrete domains of the plasma membrane. This enrichment is probably important for localization of function, but it is not clear how it is accomplished. One proposal for epithelial cells is that proteins contain targeting signals which guide preferential accumulation in basal or apical membranes. The growth-cone membrane of a neuron serves as a specialized transduction system, which helps to convert cues from its environment into regulated growth. Because it can be physically separated from the cell soma, it has been possible to show that the growth-cone membrane contains a restricted set of total cellular proteins, although, to our knowledge, no proteins are limited to that structure. One of the most prominent proteins in the growth-cone membrane is GAP-43. Basi et al. have suggested that the N-terminus of GAP-43 might be important for the binding of GAP-43 to the growth-cone membrane. Skene and Virag recently found that the cysteines in the N-terminus are fatty-acylated and that this post-translational modification correlates with membrane-binding ability. We investigated the binding of GAP-43 to the growth-cone membrane by mutational analysis and by laser-scanning confocal microscopy of fusion proteins that included regions of GAP-43 and chloramphenicol acetyltransferase (CAT). We found that a short stretch of the GAP-43 N-terminus suffices to direct accumulation in growth-cone membranes, especially in the filopodia. This supports a previous proposal for the importance of this region of GAP-43 in determining the membrane distribution of GAP-43.

Adrenal Gland Neoplasms↗
PubMed1989

The neuronal growth-associated protein GAP-43 induces filopodia in non-neuronal cells.

The neuron-specific protein GAP-43 is associated with the membrane of the nerve growth cone and thus may be important to the activity of this distinctive neuronal structure. Transient transfection of COS and NIH 3T3 cells with appropriate vectors resulted in expression of GAP-43 in these non-neuronal cells; as in neurons, transfected GAP-43 associated with the membrane. In addition, many long fine filopodial processes extended from the periphery of such transfected cells. Stable CHO cell lines expressing GAP-43 also exhibited processes that were more numerous, far longer, and more complex than those of CHO cell lines not transfected or transfected with control plasmids. Thus GAP-43 may directly contribute to growth cone activity by regulating cell membrane structure and enhancing extension of filopodial processes.

Animals↗
PubMed1989

GAP-43 gene expression during development: persistence in a distinctive set of neurons in the mature central nervous system.

GAP-43 is a rapidly transported axonal protein most prominently expressed in regenerating and developing nerves. However, the low level persistence of GAP-43 in the adult CNS where growth and regenerative capacity are minimal may additionally indicate a role for this molecule in neuronal remodeling. Previous studies have revealed GAP-43 immunoreactivity in neurites throughout many regions of the CNS. To identify the CNS neurons that express GAP-43 at different stages of development, we utilized in situ hybridization and immunocytochemistry; the latter was performed with an antibody that recognizes GAP-43 immunoreactivity in both perikarya and neurites. In the perinatal period GAP-43 is expressed in all neurons. Subsequently its expression becomes progressively restricted such that by maturity most neurons no longer express detectable levels, although GAP-43 expression is still moderately high in the adult entorhinal cortex, and strikingly high in the adult hippocampus and olfactory bulb. In light of current notions about the function of GAP-43, it is tempting to speculate that this anatomy denotes neurons engaged in structural remodeling and functional plasticity.

Aging↗
PubMed1989

A Drosophila gene expressed in the embryonic CNS shares one conserved domain with the mammalian GAP-43.

By cross hybridization with the mammalian growth-related protein, GAP-43, we have isolated several Drosophila cDNAs and genomic sequences. These sequences correspond to a single copy gene that encodes two developmentally regulated transcripts 2.4 and 2.0 kb in length. The predicted protein sequence from the cDNAs contains a stretch of 20 amino acids closely related to the mammalian GAP-43 protein. These residues are also highly conserved in a cDNA isolated from the nematode C. elegans. Prior to dorsal closure, expression of the Drosophila gene is observed in non-neuronal tissues, especially in the mesectoderm and presumptive epidermis, both in a metameric pattern. After dorsal closure, expression becomes restricted to sets of cells that are segmentally reiterated along the periphery of the nervous system. These cells appear to include at least one specific set of glia that may establish scaffolding for the development of the longitudinal neuropile.

Amino Acid Sequence↗
PubMed1988

Dual regulation of GAP-43 gene expression by nerve growth factor and glucocorticoids.

GAP-43 is a neural-specific protein that is believed integral to neurite growth and to the plasticity of neuronal structure. Its gene expression is regulated in vivo and correlates with periods of axonal growth. We investigated the regulation of GAP-43 gene expression in PC12 cells, which are believed to resemble precursor cells of the adrenomedullary lineage. In these cells, nerve growth factor (NGF) increases GAP-43 expression, and corticosteroids decrease it. Corticosteroids diminish GAP-43 levels even in cells already differentiated by NGF, as well as in primary neurons of the superior cervical ganglion. Neither the NGF nor the steroid effect requires new protein synthesis. Nuclear run-on experiments show that the steroid repression is mediated at the level of gene transcription but that the NGF effect is likely to be posttranscriptional. NGF and corticosteroids are known to regulate bimodally the cell fate decision of sympathoadrenal precursors, with NGF promoting the neuronal phenotype and steroids promoting the chromaffin phenotype. The regulation of GAP-43 is consistent with the notion that this gene is bimodally regulated during these cell fate decisions.

Adrenal Gland Neoplasms↗
PubMed1988

Cloning of human GAP-43: growth association and ischemic resurgence.

GAP-43 is a growth cone protein expressed in neurons especially during periods of axonal elongation. Poor repair in the adult mammalian CNS has been ascribed to restraints upon its expression. We have cloned human GAP-43 cDNA to investigate its potential involvement in neurological illness. Analysis of postmortem human brain tissue disclosed uniformly high expression of GAP-43 throughout the neonatal brain, whereas in the adult brain high levels of GAP-43 persist only in discrete regions. However, in the wake of ischemic injury in the adult brain, regions normally low in GAP-43 reexpress it at high levels, suggesting a role for GAP-43 in remodeling and repair of mature CNS neurons.

Adolescent↗
PubMed1987

Cloning of complementary DNA for GAP-43, a neuronal growth-related protein.

GAP-43 is one of a small subset of cellular proteins selectively transported by a neuron to its terminals. Its enrichment in growth cones and its increased levels in developing or regenerating neurons suggest that it has an important role in neurite growth. A complementary DNA (cDNA) that encodes rat GAP-43 has been isolated to study its structural characteristics and regulation. The predicted molecular size is 24 kilodaltons, although its migration in SDS-polyacrylamide gels is anomalously retarded. Expression of GAP-43 is limited to the nervous system, where its levels are highest during periods of neurite outgrowth. Nerve growth factor or adenosine 3',5'-monophosphate induction of neurites from PC12 cells is accompanied by increased GAP-43 expression. GAP-43 RNA is easily detectable, although at diminished levels, in the adult rat nervous system. This regulation of GAP-43 is concordant with a role in growth-related processes of the neuron, processes that may continue in the mature animal.

Amino Acid Sequence↗
PubMed1987

Regulation of tyrosine hydroxylase gene expression during differentiation of neuroblastoma cells.

Differentiation of N1E-115 neuroblastoma cells into neuron-like cells, with extension of neurites and acquisition of excitable membranes, can be induced by dimethyl sulfoxide (DMSO). We have found this differentiation to be accompanied by an increase in tyrosine hydroxylase (TH) mRNA, an increase disproportionate to changes in mRNAs for other measured, non-neuron-specific genes. The mRNA increases slowly over several days and falls gradually after removal of DMSO. Nuclear run-on studies suggest that a change in the rate of transcription cannot explain the increase in steady-state mRNA levels. TH mRNA half-life does, however, increase. This suggests that regulation is exerted in this case not at the level of transcription but rather at that of mRNA stability.

Animals↗
PubMed1987

Neurochemical characteristics of a novel dorsal root ganglion X neuroblastoma hybrid cell line, F-11.

We investigated the properties of the novel dorsal root ganglion (DRG) hybrid cell line F-11 to see how closely these cells resembled normal DRG cells. Under normal growth conditions, F-11 cells appeared to contain several short neurite-like processes. However, these cells could also be grown under conditions in which they showed a much more extensive neuronal morphology, exhibiting many long neurites. Several differentiated features of DRG cells were present on F-11 cells. These included the presence of delta-opioid receptors, receptors for prostaglandins and bradykinin, and dihydropyridine-sensitive calcium channels. F-11 cells also synthesized and released a substance P-like compound, as determined by immunoreactivity. Both the number of bradykinin receptors and the voltage-sensitive calcium influx increased on cell differentiation. Opioid agonists (delta-specificity) were found to decrease cyclic AMP levels in F-11 cells in a naloxone- and pertussis toxin-reversible fashion. Bradykinin stimulated the synthesis of inositol-1,4-bisphosphate and inositol-1,4,5-trisphosphate. Ca2+ channel agonists stimulated voltage-sensitive Ca2+ influx in a dose-dependent, stereospecific manner, whereas Ca2+ channel antagonists inhibited Ca2+ influx. F-11 cells should, therefore, prove useful as models for authentic DRG neurons.

Animals↗
PubMed1987

Recognition of specific targets by cultured dorsal root ganglion neurons.

We have assessed the effects of different target cell populations on axonally transported proteins by the use of compartmental cell culture systems that separate the soma from the growing axons of rat sensory neurons. The labeling of 3 rapidly transported proteins diminishes when the growing axon contacts spinal cord cells (which are normal in vivo targets), and remains unaffected by contact with fibroblasts or heart cells. Medium conditioned by spinal cord cells does not exert this effect. Thus, specific classes of cells may be distinguished as target tissue by sensory neurons in vitro. Such recognition is accompanied by specific molecular changes in axonally transported proteins.

Animals↗