Stimulation of the pituitary-adrenocortical axis by nerve growth factor.

Nerve growth factor (NGF) is a protein essential for the development and maintenance of the peripheral sympathetic nervous system, causing responsive neurones to increase in size and to extend neurites. Biochemically, the selective induction of tyrosine hydroxylase (TH) and dopamine beta-hydroxylase key enzymes in catecholamine biosynthesis is one of its most characteristic effects. Both the morphological and biochemical effects are modulated by glucocorticoids, suggesting a close relationship between specific effects of NGF and hormone action. NGF has been shown to induce an increase in adrenal cyclic AMP in intact but not in hypophysectomised rats, and so we have looked directly at the effect of systemic administration of NGF on the hypothalamo-pituitary-adrenal axis. We report here that NGF induced an enhanced secretion of adrenocorticotropin (ACTH) and a prolonged increase in plasma glucocorticoid concentration after intravenous (i.v.) injection. Such effects could have important implications for the biological activity of NGF.     

New protein fold revealed by a 2.3-A resolution crystal structure of nerve growth factor

Nerve growth factor (NGF) is a member of an expanding family of neurotrophic factors (including brain-derived neurotrophic factor and the neurotrophins) that control the development and survival of certain neuronal populations both in the peripheral and in the central nervous systems. Its biological effects are mediated by a high-affinity ligand-receptor interaction and a tyrosine kinase signalling pathway. A potential use for NGF and its relatives in the treatment of neurological disorders such as Alzheimer's disease and Parkinson's disease requires an understanding of the structure-function relationships of NGF. NGF is a dimeric molecule, with 118 amino acids per protomer. We report the crystal structure of the murine NGF dimer at 2.3-A resolution, which reveals a novel protomer structure consisting of three antiparallel pairs of beta strands, together forming a flat surface. Two subunits associate through this surface, thus burying a total of 2,332 A. Four loop regions, which contain many of the variable residues observed between different NGF-related molecules, may determine the different receptor specificities. A clustering of positively charged side chains may provide a complementary interaction with the acidic low-affinity NGF receptor. The structure provides a model for rational design of analogues of NGF and its relatives and for testing the NGF-receptor recognition determinants critical for signal transduction.     

Engagement of the high-affinity IgE receptor activates src protein-related tyrosine kinases.

The high-affinity IgE receptor (Fc epsilon RI), which is expressed on the surface of mast cells and basophils, has a central role in immediate allergic responses. In the rat basophilic leukaemia cell line RBL-2H3, which is a model system for the analysis of Fc epsilon RI-mediated signal transduction, surface engagement of Fc epsilon RI induces histamine release and the tyrosine phosphorylation of several distinct proteins. Although the alpha, beta, and gamma subunits of Fc epsilon RI lack intrinsic tyrosine protein kinase (TPK) activity, a kinase that copurifies with Fc epsilon RI phosphorylates the beta and gamma subunits of the receptor on tyrosine residues. We report here that in RBL-2H3 cells, p56lyn and pp60c-src are activated after Fc epsilon RI crosslinking, and p56lyn coimmunoprecipitates with Fc epsilon RI. In the mouse mast-cell line PT-18, another cell type used to study FC epsilon RI-mediated signalling, tyrosine phosphorylation of proteins is also an immediate consequence of receptor crosslinking. Notably, the only detectable src protein-related TPK in PT-18 cells is p62c-yes, and it is this TPK that is activated on Fc epsilon RI engagement and coimmunoprecipitates with the receptor. Therefore, it seems that different src protein-related TPKs can associate with the same receptor and become activated after receptor engagement.     

Regulation of carbamoyl phosphate synthetase by MAP kinase

The de novo synthesis of pyrimidine nucleotides is required for mammalian cells to proliferate. The rate-limiting step in this pathway is catalysed by carbamoyl phosphate synthetase (CPS II), part of the multifunctional enzyme CAD. Here we describe the regulation of CAD by the mitogen-activated protein (MAP) kinase cascade. When phosphorylated by MAP kinase in vitro or activated by epidermal growth factor in vivo, CAD lost its feedback inhibition (which is dependent on uridine triphosphate) and became more sensitive to activation (which depends upon phosphoribosyl pyrophosphate). Both these allosteric regulatory changes favour biosynthesis of pyrimidines for growth. They were accompanied by increased epidermal growth factor-dependent phosphorylation of CAD in vivo and were prevented by inhibition of MAP kinase. Mutation of a consensus MAP kinase phosphorylation site abolished the changes in CAD allosteric regulation that were stimulated by growth factors. Finally, consistent with an effect of MAP kinase signalling on CPS II activity, epidermal growth factor increased cellular uridine triphosphate and this increase was reversed by inhibition of MAP kinase. Hence these studies may indicate a direct link between activation of the MAP kinase cascade and de novo biosynthesis of pyrimidine nucleotides.     

Immunohistochemical localization of endogenous nerve growth factor

Nerve growth factor (NGF) has been proposed as a trophic molecule essential for the development of sympathetic and primary sensory neurones. In newborn mice and rats, administration of nerve growth factor results in an increase in the number of surviving neurones, whereas administration of antiserum to NGF decreases neuronal survival. Thus it has been proposed that the factor is produced and secreted by the relevant target tissues to provide trophic support for the ingrowing nerves. The site of synthesis of nerve growth factor is still unknown, and it has been emphasized that a precise physiological role for the molecule cannot be ascribed until the cell types that produce it are known. I report here the use of immunohistochemistry to localize endogenous NGF in the rat iris, a tissue in which there is sound biochemical evidence for the production of NGF activity. Surprisingly, the results reveal that NGF can be detected readily in Schwann cells, but not in smooth muscle cells of the iris when it is sympathetically denervated or cultured.     

Alterations in the surface properties of cells responsive to nerve growth factor.

An initial effect of nerve growth factor on a responsive nerve cell line is to increase intracellular cyclic AMP, which in turn leads to a mobilisation of calcium ions. These events are correlated with structural changes in the plasma membrane, increased cell-substratum adhesion, and neurite outgrowth.     

Proliferation and differentiation of neuronal stem cells regulated by nerve growth factor

Nerve growth factor plays an important part in neuron-target interactions in the late embryonic and adult brain. We now report that this growth factor controls the proliferation of neuronal precursors in a defined culture system of cells derived from the early embryonic brain. Neuronal precursor cells were identified by expression of the intermediate filament protein nestin. These cells proliferate in response to nerve growth factor but only after they have been exposed to basic fibroblast growth factor. On withdrawal of nerve growth factor, the proliferative cells differentiate into neurons. Thus, in combination with other growth factors, nerve growth factor regulates the proliferation and terminal differentiation of neuroepithelial stem cells.     

Jelly belly protein activates the receptor tyrosine kinase Alk to specify visceral muscle pioneers

The secreted protein Jelly belly (Jeb) is required for an essential signalling event in Drosophila muscle development. In the absence of functional Jeb, visceral muscle precursors are normally specified but fail to migrate and differentiate. The structure and distribution of Jeb protein implies that Jeb functions as a signal to organize the development of visceral muscles. Here we show that the Jeb receptor is the Drosophila homologue of anaplastic lymphoma kinase (Alk), a receptor tyrosine kinase of the insulin receptor superfamily. Human ALK was originally identified as a proto-oncogene, but its normal function in mammals is not known. In Drosophila, localized Jeb activates Alk and the downstream Ras/mitogen-activated protein kinase cascade to specify a select group of visceral muscle precursors as muscle-patterning pioneers. Jeb/Alk signalling induces the myoblast fusion gene dumbfounded (duf; also known as kirre) as well as org-1, a Drosophila homologue of mammalian TBX1, in these cells.