RESUMEN
A novel member of the neuropoietic cytokine family has been cloned and the protein expressed and characterized. In an effort to identify novel secreted proteins, an algorithm incorporating neural network algorithms was applied to a large EST database. A full-length clone was identified that is 1710 bp in length and has a single open reading frame of 225 amino acids. This new cytokine is most homologous to cardiotrophin-1, having a similarity and an identity of 46 and 29%, respectively, and therefore we have named it cardiotrophin-like cytokine (CLC). Northern hybridization analysis identified a 1.4-kb messenger RNA that is highly expressed in spleen and peripheral leukocytes. Purified recombinant CLC induced the activation of NFkappaB and SRE reporter constructs in the TF-1, U937, and M1 cell lines. Furthermore, the signal transduction pathway for CLC was characterized in the neuroblastoma cell line SK-N-MC and found to involve tyrosine phosphorylation of gp130 and STAT-1.
Asunto(s)
Citocinas/genética , Bases de Datos Factuales , Etiquetas de Secuencia Expresada , Redes Neurales de la Computación , Secuencia de Aminoácidos , Antígenos CD/metabolismo , Línea Celular , Cromosomas Humanos Par 11/genética , Clonación Molecular , Receptor gp130 de Citocinas , Citocinas/química , Citocinas/aislamiento & purificación , Citocinas/farmacología , Proteínas de Unión al ADN/metabolismo , Regulación de la Expresión Génica , Genes Reporteros , Humanos , Glicoproteínas de Membrana/metabolismo , Datos de Secuencia Molecular , Fosforilación/efectos de los fármacos , Fosfotirosina/metabolismo , Filogenia , ARN Mensajero/análisis , ARN Mensajero/genética , Factor de Transcripción STAT1 , Homología de Secuencia de Aminoácido , Transducción de Señal/efectos de los fármacos , Transactivadores/metabolismoRESUMEN
Myeloid progenitor inhibitory factor (MPIF)-2 is a beta-chemokine with select and potent activities on eosinophils and myeloid progenitors. In the beta-chemokine family, biological activity is modulated by differential processing of the amino-terminus. Here, for MPIF-2, we describe the biological activities of NH(2)-terminal deletion mutants and compare regions necessary for eosinophil and myeloid progenitor activities. Five MPIF-2 proteins with deletions at the amino-terminus were produced in Escherichia coli and assayed for calcium mobilization, chemotaxis and receptor binding activities on eosinophils, and for their ability to inhibit colony formation of human myeloid bone marrow progenitors. For eosinophils, deletion of the first two amino acids did not markedly alter activity, while subsequent truncations result in a complete loss of activity. One of the MPIF-2 mutants, MPIF-2 (P30-R99) was converted from an agonist to an antagonist of eotaxin, MPIF-2 and MCP-4 functional responses in eosinophil calcium flux and chemotaxis assays. Surprisingly, while displaying a complete loss of agonist activity toward eosinophils, MPIF-2 (P30-R99) retains ability to inhibit human bone marrow myeloid progenitor cell colony formation. In addition, processing at the amino terminus of MPIF-2 in vivo, may result in a chemokine with altered biological activities.