An intrinsic guanine nucleotide exchange inhibitor in Gi2 alpha. Significance of G-protein self-suppression which antagonizes receptor signal.

The alpha subunit of Gi2 (Gi2 alpha) is a member of the heterotrimeric G protein family, which transduces receptor signals as a proto-oncogene product. We have found a novel self-suppressive region in Gi2 alpha near its C terminus. A polypeptide consisting of residues 338-352 of Gi2 alpha (Gi2 alpha-339-352) antagonizes receptor- and receptor peptide-stimulated Gi2 alpha activation, without affecting basal activity.

Activated mutants of the alpha subunit of G(o) promote an increased number of neurites per cell.

The high concentration of the GTP-binding protein G(o) in the neuronal growth cone suggests that G(o) activation state may after neurite outgrowth. We find that activation of pertussis toxin-sensitive G-proteins by mastoparan increases neurite outgrowth from neuroblastoma cells. To examine G(o) activation specifically, point mutations homologous to activating, oncogenic mutations in alpha i2 and alpha s were introduced into the alpha subunit of G(o).

An amino-terminal domain of the growth-associated protein GAP-43 mediates its effects on filopodial formation and cell spreading.

GAP-43 is a neuronal protein that is believed to be important to neuronal growth and nerve terminal plasticity. It is enriched on the inner surface of growth cone membranes, a localization that may depend upon palmitoylation of Cys3 and Cys4. It is a major substrate for protein kinase C, which phosphorylates Ser41.

GAP-43 augments G protein-coupled receptor transduction in Xenopus laevis oocytes.

The neuronal protein GAP-43 is thought to play a role in determining growth-cone motility, perhaps as an intracellular regulator of signal transduction, but its molecular mechanism of action has remained unclear. We find that GAP-43, when microinjected into Xenopus laevis oocytes, increases the oocyte response to G protein-coupled receptor agonists by 10- to 100-fold. Higher levels of GAP-43 cause a transient current flow, even without receptor stimulation.

Functional expression of sodium channel mutations identified in families with periodic paralysis.

Two mutations in the sodium channel alpha subunit that have been implicated as the cause of periodic paralysis were studied by functional expression in a mammalian cell line. Both mutations disrupted inactivation without affecting the time course of the onset of the sodium current or the single-channel conductance. This is the same functional defect that was observed in myotubes cultured from affected patients and proves that these mutations are not benign polymorphisms.

Mediation by G proteins of signals that cause collapse of growth cones.

During development, motion of nerve growth cones ceases on contact with particular targets. The signaling mechanism is unknown. In culture, growth cone collapse can be caused by solubilized embryonic brain membranes, central nervous system myelin, a 35-kilodalton protein isolated from myelin, and mastoparan.

GAP-43 as a plasticity protein in neuronal form and repair.

Neurons exhibit a remarkable plasticity of form, both during neural development and during the subsequent remodelling of synaptic connectivity. Here we review work on GAP-43 and G0, and focus upon the thesis that their interaction may endow neurons with such plasticity. We also present new data on the role of G proteins in neurite growth, and on the interaction of GAP-43 and actin.

Palmitoylation alters protein activity: blockade of G(o) stimulation by GAP-43.

The addition of palmitate to cysteine residues enhances the hydrophobicity of proteins, and consequently their membrane association. Here we have investigated whether this type of fatty acylation also regulates protein-protein interactions. GAP-43 is a neuronal protein that increases guanine nucleotide exchange by heterotrimeric G proteins.

GAP-43 as a modulator of G protein transduction in the growth cone.

Much circumstantial evidence that GAP-43 is involved in neuronal growth cone function has accumulated over the last ten years. The expression of the protein is closely correlated both temporally and spatially with periods of axonal outgrowth, and the protein is highly concentrated in the growth cone membrane. There is direct evidence that overexpression of the protein can alter cell shape.

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.

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.

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).

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.

Localization of angiotensin converting enzyme in the ciliary epithelium of the rat eye.

Angiotensin converting enzyme (ACE, E.C. 3.

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.

Characterization of a neutral, divalent cation-sensitive endopeptidase: a possible role in neuropeptide processing.

A trypsin-like endopeptidase which cleaves the synthetic substrate Dansyl-Phe-Leu-Arg-Arg-Ala-Ser-Leu-Gly-COOH (Dansyl-Phe-Kemptide) primarily at the Arg4-Ala5 bond has been partially purified from bovine adrenal chromaffin granules, brain and liver. The enzyme appears to have a relatively homogeneous tissue distribution, although highest levels were found in brain regions such as the hippocampus and corpus striatum. Sucrose density gradient fractionation established that enzyme activity assayed at pH 8.

Enkephalin convertase: characterization and localization using [3H]guanidinoethylmercaptosuccinic acid.

Enkephalin convertase (carboxypeptidase E,H; EC 3.4.17.

Enkephalin convertase in the gastrointestinal tract an associated organs characterized and localized with [3H]guanidinoethylmercaptosuccinic acid.

Enkephalin convertase (carboxypeptidase EH; EC 3.4.17.

Angiotensin converting enzyme immunohistochemistry in rat brain and pituitary gland: correlation of isozyme type with cellular localization.

We have localized angiotensin converting enzyme in rat brain and pituitary gland immunohistochemically with an anti-rat lung angiotensin converting enzyme monoclonal antibody. The distribution of immunoreactive angiotensin converting enzyme is identical with that of binding sites for the angiotensin converting enzyme inhibitor, [3H]captopril. Most intense staining is in the choroid plexus and subfornical organ, with intermediate values in the caudate-putamen, globus pallidus, entopeduncular nucleus, pars reticulata of the substantia nigra, posterior pituitary and anterior pituitary.

Morphological and physiological survival of goldfish Mauthner axons isolated from their somata by spinal cord crush.

Axon segments isolated from their somata degenerate within days or months depending on species and neuronal type. To better understand the time course of morphological and physiological changes associated with degeneration of axon segments of vertebrate central neurons, we have studied the goldfish Mauthner axon (M-axon) when it has been separated from its soma by spinal cord crush. M-axon segments survive morphologically for at least 77 days at 14 degrees C.

Differential ontogeny of rat brain peptidases: prenatal expression of enkephalin convertase and postnatal development of angiotensin-converting enzyme.

We quantitated the levels of two peptidases in the developing rat brain as a means to determine their function. Enkephalin convertase (EC 3.4.

Parkinson's disease: nigral receptor changes support peptidergic role in nigrostriatal modulation.

Autoradiographic studies reveal densities of binding to somatostatin, neurotensin, mu-opiate, and benzodiazepine receptors in substantia nigra specimens from neurologically normal human brains. Binding to nigral angiotensin converting enzyme is also dense, whereas more modest densities of kappa-opiate, dopamine, and serotonin receptors are noted. In nigral specimens taken from patients with idiopathic Parkinson's disease, substantial reductions in somatostatin, neurotensin, mu-opiate and kappa-opiate receptors contrast with more modest reductions in dopamine and benzodiazepine I receptor subtypes.

Enkephalin convertase: localization to specific neuronal pathways.

3H-Guanidinoethylmercaptosuccinic acid (GEMSA) selectively labels the carboxypeptidase B-like enzyme enkephalin convertase (EC) in rat brain tissue sections. We have used autoradiography with 3H-GEMSA to map membrane-bound EC in the rat forebrain and, in conjunction with lesioning techniques, to localize EC to specific neuronal pathways. The highest levels of EC are in the median eminence.

Angiotensin-converting enzyme localized in the rat pituitary and adrenal glands by [3H]captopril autoradiography.

We have localized angiotensin-converting enzyme (EC 3.14.5.

Characterization of angiotensin converting enzyme by [3H]captopril binding.

We demonstrate that [3H]captopril selectively labels angiotensin converting enzyme (EC 3.14.15.