RESUMO
Endothelial monocyte-activating polypeptide II (EMAP II) is a proinflammatory cytokine and a chemoattractant for leukocytes. The mature cytokine is formed in apoptotic cells by cleavage of the precursor proEMAP II. Here we show that caspase-7 is capable of cleaving proEMAP II in vitro. A proEMAP II mutant, in which the ASTD cleavage site was changed to the sequence ASTA, was not processed by caspase-7. The caspase-7-mediated generation and release of mature EMAP II may provide a mechanism for leukocyte recruitment to sites of programmed cell death, and thus may link apoptosis to inflammation.
Assuntos
Caspases/metabolismo , Citocinas , Proteínas de Neoplasias/metabolismo , Proteínas de Ligação a RNA/metabolismo , Sequência de Aminoácidos , Animais , Apoptose , Sequência de Bases , Sítios de Ligação/genética , Caspase 3 , Caspase 7 , Caspases/genética , Primers do DNA/genética , Técnicas In Vitro , Mediadores da Inflamação/metabolismo , Cinética , Camundongos , Mutagênese Sítio-Dirigida , Proteínas de Neoplasias/genética , Precursores de Proteínas/genética , Precursores de Proteínas/metabolismo , Processamento de Proteína Pós-Traducional , Proteínas de Ligação a RNA/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Especificidade por SubstratoRESUMO
Endothelial monocyte-activating polypeptide II (EMAP II) is a chemoattractant for monocytes and granulocytes. EMAP II is translated as a precursor protein, proEMAP II, and is proteolytically cleaved to become the mature, biologically active cytokine. In this study we show that the EMAP II mRNA and the EMAP II precursor protein are constitutively expressed by all cell types analyzed in vitro, whereas the mature cytokine is only present in the supernatant of apoptotic cells. During mouse embryogenesis we found widespread expression of the EMAP II mRNA with transcripts being abundant in areas of tissue remodeling, where a large number of apoptotic cells could be detected by TUNEL staining. In the adult mouse, strong expression of the EMAP II mRNA is restricted to the brain, testis and thymus. Interestingly, prominent signals for EMAP II mRNA are found in local correlation with sites of apoptosis in thymus and testis but not in the brain. We propose that during development, the generation and release of the mature EMAP II may provide a mechanism for the recruitment of phagocytic cells to sites of programmed cell death. In the adult brain, the generation of mature EMAP II may contribute to the recruitment of monocytes and the immunosurveillance of this tissue.
Assuntos
Apoptose/fisiologia , Citocinas , Inibidores do Crescimento/metabolismo , Camundongos/embriologia , Proteínas de Neoplasias/metabolismo , Precursores de Proteínas/metabolismo , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/metabolismo , Animais , Aorta/citologia , Northern Blotting , Western Blotting , Encéfalo/citologia , Encéfalo/metabolismo , Bovinos , Células Cultivadas , Meios de Cultura Livres de Soro , Embrião de Mamíferos/metabolismo , Endotélio Vascular/citologia , Inibidores do Crescimento/genética , Humanos , Hibridização In Situ , Marcação In Situ das Extremidades Cortadas , Rim/citologia , Rim/metabolismo , Masculino , Camundongos/crescimento & desenvolvimento , Proteínas de Neoplasias/genética , Precursores de Proteínas/genética , Proteínas de Ligação a RNA/genética , Testículo/citologia , Testículo/metabolismo , Timo/citologia , Timo/metabolismoRESUMO
Endothelial monocyte-activating polypeptide II (EMAP II) is a proinflammatory cytokine and a chemoattractant for monocytes. We show here that, in the mouse embryo, EMAP II mRNA was most abundant at sites of tissue remodeling where many apoptotic cells could be detected by terminal deoxynucleotidyltransferase-mediated dUTP end labeling. Removal of dead cells is known to require macrophages, and these were found to colocalize with areas of EMAP II mRNA expression and programmed cell death. In cultured cells, post-translational processing of pro-EMAP II protein to the mature released EMAP II form (23 kDa) occurred coincidentally with apoptosis. Cleavage of pro-EMAP II could be abrogated in cultured cells by using a peptide-based inhibitor, which competes with the ASTD cleavage site of pro-EMAP II. Our results suggest that the coordinate program of cell death includes activation of a caspase-like activity that initiates the processing of a cytokine responsible for macrophage attraction to the sites of apoptosis.