RESUMO
Bcl-2 proteins are acknowledged as key regulators of programmed cell death. However, increasing data suggest additional roles, including regulation of the cell cycle, metabolism and cytoskeletal dynamics. Here we report the discovery and characterization of a new Bcl-2-related multidomain apoptosis accelerator, Bcl-wav, found in fish and frogs. Genetic and molecular studies in zebrafish indicate that Bcl-wav and the recently identified mitochondrial calcium uniporter (MCU) contribute to the formation of the notochord axis by controlling blastomere convergence and extension movements during gastrulation. Furthermore, we found that Bcl-wav controls intracellular Ca(2+) trafficking by acting on the mitochondrial voltage-dependent anion channel, and possibly on MCU, with direct consequences on actin microfilament dynamics and blastomere migration guidance. Thus, from an evolutionary point of view, the original function of Bcl-2 proteins might have been to contribute in controlling the global positioning system of blastomeres during gastrulation, a critical step in metazoan development.
Assuntos
Canais de Cálcio/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Canal de Ânion 1 Dependente de Voltagem/metabolismo , Peixe-Zebra/embriologia , Actinas/genética , Sequência de Aminoácidos , Animais , Apoptose/genética , Transporte Biológico/genética , Blastômeros/metabolismo , Cálcio/metabolismo , Canais de Cálcio/genética , Movimento Celular/genética , Células Cultivadas , Gástrula/embriologia , Gastrulação/genética , Técnicas de Silenciamento de Genes , Proteínas de Fluorescência Verde/genética , Células HeLa , Humanos , Camundongos , Mitocôndrias/metabolismo , Dados de Sequência Molecular , Morfogênese , Morfolinos/genética , Notocorda/metabolismo , Alinhamento de Sequência , Análise de Sequência de DNA , Canal de Ânion 1 Dependente de Voltagem/genéticaRESUMO
BACKGROUND: Neonatal alloimmune thrombocytopenia is mostly due to the presence of maternal antibodies against the fetal platelet antigen HPA-1a on the platelet integrin GPIIb-IIIa. Accurate detection of anti-HPA-1a antibodies in the mother is, therefore, critical. Current diagnostic assays rely on the availability of pools of human platelets that vary according to donors and blood centers. There is still no satisfactory standardization of these assays. DESIGN AND METHODS: Peptide aptamer was used to detect and identify HPA-1a-specific antibodies in human serum that do not require human platelets. A peptide aptamer library was screened using an anti-HPA-1a human monoclonal antibody as a bait to isolate an aptamer that mimics the human platelet antigen HPA-1a. RESULTS: This is the first report in platelet immunology of the use of a peptide aptamer for diagnostic purposes. This assay gives better results than the MAIPA currently in use, detecting around 90% of the expected alloantibodies. CONCLUSIONS: This assay could help define a standard for the quantitation of anti-HPA antibodies. This report also demonstrates that peptide aptamers can potentially detect a variety of biomarkers in body fluids; this is of particular interest for diagnostic purposes.
Assuntos
Anticorpos Anti-Idiotípicos/sangue , Antígenos de Plaquetas Humanas/imunologia , Aptâmeros de Nucleotídeos , Plaquetas/imunologia , Isoanticorpos/sangue , Fragmentos de Peptídeos , Complexo Glicoproteico GPIIb-IIIa de Plaquetas/imunologia , Trombocitopenia Neonatal Aloimune/diagnóstico , Sequência de Aminoácidos , Anticorpos Anti-Idiotípicos/imunologia , Anticorpos Monoclonais/imunologia , Complexo Antígeno-Anticorpo , Western Blotting , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Imunoensaio , Imunoprecipitação , Recém-Nascido , Integrina beta3 , Isoanticorpos/imunologia , Dados de Sequência Molecular , Trombocitopenia Neonatal Aloimune/sangue , Trombocitopenia Neonatal Aloimune/imunologiaRESUMO
Bcl-2 family members are key regulators of apoptosis. Their involvement in other cellular processes has been so far overlooked. We have studied the role of the Bcl-2 homolog Nrz in the developing zebrafish. Nrz was found to be localized to the yolk syncytial layer, a region containing numerous mitochondria and ER membranes. Nrz knockdown resulted in developmental arrest before gastrulation, due to free Ca(2+) increase in the yolk cell, activating myosin light chain kinase, which led to premature contraction of actin-myosin cables in the margin and separation of the blastomeres from the yolk cell. In the yolk syncytial layer, Nrz appears to prevent the release of Ca(2+) from the endoplasmic reticulum by directly interacting with the IP3R1 Ca(2+) channel. Thus, the Bcl-2 family may participate in early development, not only by controlling apoptosis but also by acting on cytoskeletal dynamics and cell movements via Ca(2+) fluxes inside the embryo.
