RESUMO
The ErbB4 receptor tyrosine kinase possesses both tumour suppressor and oncogenic activities. Thus pharmacological agents are needed to help elucidate ErbB4 functions. However, limitations of existing ErbB4 agonists and antagonists have led us to seek novel ErbB4 antagonists. The Q43L mutant of the ErbB4 agonist NRG2ß (neuregulin 2ß) stimulates ErbB4 tyrosine phosphorylation, yet fails to stimulate ErbB4 coupling to cell proliferation. Thus in the present paper we hypothesize that NRG2ß/Q43L may be an ErbB4 antagonist. NRG2ß/Q43L competitively antagonizes agonist stimulation of ErbB4 coupling to cell proliferation. NRG2ß/Q43L stimulates less ErbB4 tyrosine phosphorylation than does NRG2ß. In addition, NRG2ß stimulation of cell proliferation requires PI3K (phosphoinositide 3-kinase) activity and NRG2ß stimulates greater Akt phosphorylation than does NRG2ß/Q43L. Moreover, EGFR [EGF (epidermal growth factor) receptor] kinase activity (but not that of ErbB4) is critical for coupling ErbB4 to proliferation. Experiments utilizing ErbB4 splicing isoforms and mutants suggest that NRG2ß and NRG2ß/Q43L may differentially stimulate ErbB4 coupling to the transcriptional co-regulator YAP (Yes-associated protein). Finally, NRG2ß/Q43L competitively antagonizes agonist stimulation of EGFR and ErbB2/ErbB3, indicating that NRG2ß/Q43L is a pan-ErbB antagonist. Thus we postulate that NRG2ß/Q43L and other antagonistic ligands stimulate ErbB tyrosine phosphorylation on a set of residues distinct from that stimulated by agonists, thus suggesting a novel mechanism of ErbB receptor regulation. Moreover, NRG2ß/Q43L and related ligand-based antagonists establish a paradigm for the discovery of anti-ErbB therapeutics.
Assuntos
Receptores ErbB/antagonistas & inibidores , Mutação de Sentido Incorreto , Fatores de Crescimento Neural/genética , Sequência de Aminoácidos , Ligação Competitiva , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Receptores ErbB/agonistas , Receptores ErbB/metabolismo , Humanos , Isoenzimas/antagonistas & inibidores , Isoenzimas/metabolismo , Dados de Sequência Molecular , Fatores de Crescimento Neural/metabolismo , Fatores de Crescimento Neural/farmacologia , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação/efeitos dos fármacos , Domínios e Motivos de Interação entre Proteínas , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptor ErbB-2/metabolismo , Receptor ErbB-3/antagonistas & inibidores , Receptor ErbB-3/metabolismo , Receptor ErbB-4 , Transdução de SinaisRESUMO
The EGF family hormone NRG2beta potently stimulates ErbB4 tyrosine phosphorylation and coupling to IL3 independence. In contrast, the NRG2alpha splicing isoform has lower affinity for ErbB4, does not potently stimulate ErbB4 phosphorylation, and fails to stimulate ErbB4 coupling. Here we investigate these differences. The NRG2beta Q43L mutant potently stimulates ErbB4 phosphorylation but not ErbB4 coupling to IL3 independence. This failure to stimulate ErbB4 coupling is not due to differential ligand purity, glycosylation, or stability. The NRG2alpha K45F mutant potently stimulates ErbB4 phosphorylation but not ErbB4 coupling to IL3 independence. Thus, this failure to stimulate ErbB4 coupling is not due to inadequate affinity for ErbB4. In contrast, the NRG2alpha L43Q/K45F mutant stimulates ErbB4 coupling, even though it does not have greater affinity for ErbB4 than does NRG2alpha/K45F. Collectively, these data indicate that Gln43 of NRG2beta is both necessary and sufficient for NRG2 stimulation of ErbB4 coupling to IL3 independence.
Assuntos
Receptores ErbB/metabolismo , Interleucina-3/metabolismo , Fatores de Crescimento Neural/metabolismo , Processamento de Proteína , Sequência de Aminoácidos , Linhagem Celular , Receptores ErbB/agonistas , Glicina/genética , Glicina/metabolismo , Humanos , Dados de Sequência Molecular , Mutação , Fatores de Crescimento Neural/genética , Fatores de Crescimento Neural/farmacologia , Fosforilação , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Receptor ErbB-4 , Proteínas Recombinantes/farmacologia , Tirosina/metabolismoRESUMO
We re-engineered the immunoglobulin rearrangements from clonally expanded CSF B cells of three Multiple Sclerosis patients as Fab fragments, and used three methods to test for their antigen (Ag) specificity. Nine out of ten Fab fragments were reactive to Myelin Basic Protein (MBP). The one Fab that did not react to MBP was a product of receptor editing. Two of the nine MBP reactive Fabs were also reactive to GFAP and CNPase, indicating that these clones were polyreactive. Targeting the mechanisms that allow these self-reactive B cells to reside in the CSF of MS patients may prove to be a potent immunotherapeutic strategy.
Assuntos
Autoantígenos/imunologia , Linfócitos B/imunologia , Fragmentos Fab das Imunoglobulinas/imunologia , Esclerose Múltipla Recidivante-Remitente/líquido cefalorraquidiano , Esclerose Múltipla Recidivante-Remitente/imunologia , Proteína Básica da Mielina/imunologia , Células Clonais/imunologia , Células Clonais/metabolismo , Ensaio de Imunoadsorção Enzimática/métodos , Citometria de Fluxo , Rearranjo Gênico do Linfócito B/fisiologia , Genes de Imunoglobulinas , Proteína Glial Fibrilar Ácida/metabolismo , Humanos , Fragmentos Fab das Imunoglobulinas/metabolismo , Imunoprecipitação , Modelos ImunológicosRESUMO
The neuregulins (NRGs) are members of the epidermal growth factor (EGF) family of peptide growth factors. These hormones are agonists for the ErbB family of receptor tyrosine kinases, a family that includes the epidermal growth factor receptor (EGFR/ErbB1), ErbB2/Neu/HER2, ErbB3/HER3, and ErbB4/HER4. We recently observed that the EGF family hormone NRG2beta is a potent agonist for ErbB4. In contrast, NRG2alpha, a splicing isoform of the same gene that encodes NRG2beta, is a poor ErbB4 agonist. We hypothesized that carboxyl-terminal residues of NRG2beta are critical for stimulation of ErbB4 tyrosine phosphorylation and coupling to downstream signaling events. Here, we demonstrate that the substitution of a lysine residue for Phe45 in NRG2beta results in reduced ligand potency. We also demonstrate that substitution of a phenylalanine for Lys45 in NRG2alpha results in increased ligand potency. Finally, analyses of the gain-of-function NRG2alpha Chg5 mutant demonstrate that Gln43, Met47, Asn49, and Phe50 regulate ligand efficacy. Thus, these data indicate that carboxyl-terminal residues of NRG2beta are critical for activation of ErbB4 signaling. Moreover, these NRG2alpha and NRG2beta mutants reveal new insights into models for ligand-induced ErbB family receptor tyrosine phosphorylation and coupling to downstream signaling events.
Assuntos
Receptores ErbB/fisiologia , Fatores de Crescimento Neural/fisiologia , Transdução de Sinais/fisiologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Linhagem Celular , Primers do DNA , Dimerização , Dados de Sequência Molecular , Fatores de Crescimento Neural/química , Receptor ErbB-4RESUMO
During the last decade, several novel members of the Epidermal Growth Factor family of peptide growth factors have been identified. Most prominent among these are the Neuregulins or Heregulins. To date, four different Neuregulin genes have been identified (Neuregulin1-4) and several different splicing isoforms have been identified for at least two of these genes (Neuregulin1 and Neuregulin2). While Neuregulin1 isoforms have been extensively studied, comparatively little is known about Neuregulin3, Neuregulin4, or the Neuregulin2 isoforms. Indeed, there has been no systematic comparison of the activities of these molecules. Here we demonstrate that Neuregulin2alpha and Neuregulin2beta stimulate ErbB3 tyrosine phosphorylation and coupling to biological responses. In contrast, Neuregulin3 and Neuregulin4 fail to activate ErbB3 signaling. Furthermore, Neuregulin2beta, but not Neuregulin2alpha, stimulates ErbB4 tyrosine phosphorylation and coupling to biological responses. Finally, both Neuregulin3 and Neuregulin4 stimulate modest amounts of ErbB4 tyrosine phosphorylation. However, whereas Neuregulin3 stimulates a modest amount of ErbB4 coupling to biological responses, Neuregulin4 fails to stimulate ErbB4 coupling to biological responses. This suggests that there are qualitative as well as quantitative differences in ErbB family receptor activation by Neuregulin isoforms.
Assuntos
Receptores ErbB/genética , Regulação da Expressão Gênica , Genes erbB , Neurregulinas/fisiologia , Animais , Sequência de Bases , Linhagem Celular , Clonagem Molecular , Primers do DNA , Drosophila melanogaster , Regulação da Expressão Gênica/fisiologia , Neurregulinas/genética , Isoformas de Proteínas/genética , Isoformas de Proteínas/fisiologia , Ratos , Receptores Proteína Tirosina Quinases/genética , Proteínas Recombinantes/metabolismo , TransfecçãoRESUMO
Female undergraduates were divided into groups based on their rated body dissatisfaction (n = 45 operationally defined as Satisfied, n = 45 as Dissatisfied). These groups were then randomly assigned to one of three magazine categories: fashion, fitness-and-health, and news. Measures of Body Satisfaction, Depression, Anxiety, Self-esteem, Fear of Fat. Eating Attitudes, and Control of Weight were taken. Significant group main effects were found on Depression, Trait Anxiety, Eating Attitudes, Fear of Fat, and Self-esteem. A significant main effect for media was found for scores on Body Satisfaction, with the fitness-and-health Dissatisfied group reporting decreased body satisfaction following magazine exposure. No interactions were found. It appears women who are dissatisfied with their bodies may be at risk for a further decrease in body satisfaction after even a 15-min. exposure to fitness and health magazines but further follow-up measures were not made.
Assuntos
Meios de Comunicação de Massa , Satisfação Pessoal , Estimulação Luminosa , Editoração , Transtornos Somatoformes/epidemiologia , Estudantes/estatística & dados numéricos , Adulto , Imagem Corporal , Feminino , Humanos , Transtornos Somatoformes/diagnósticoRESUMO
We identified a unique antibody gene mutation pattern (i.e. "signature") in cerebrospinal fluid (CSF) B cells from multiple sclerosis (MS) patients not present in control populations. Prevalence of the signature in CSF B cells of patients at risk to develop MS predicted conversion to MS with 91% accuracy in a small cohort of clinically isolated syndrome patients. If confirmed, signature prevalence would be a novel genetic diagnostic tool candidate for patients with early demyelinating disease of the central nervous system.