RESUMO
We used DNA microarray screening to identify Ckap2 (cytoskeleton associated protein 2) as a novel p53 target gene in a mouse erythroleukemia cell line. DNA damage induces human and mouse CKAP2 expression in a p53-dependent manner and p53 activates the Ckap2 promoter. Overexpressed Ckap2 colocalizes with and stabilizes microtubules. In p53-null cells, overexpression of Ckap2 induces tetraploidy with aberrant centrosome numbers, suggesting disturbed mitosis and cytokinesis. In p53-competent cells, Ckap2 does not induce tetraploidy but activates p53-mediated cell cycle arrest and apoptosis. Our data suggest the existence of a functional positive feedback loop in which Ckap2 activates the G1 tetraploidy checkpoint and prevents aneuploidy.
Assuntos
Aneuploidia , Proteínas do Citoesqueleto/genética , Leucemia Eritroblástica Aguda/genética , Proteína Supressora de Tumor p53/genética , Animais , Apoptose/genética , Ciclo Celular/genética , Centrossomo/fisiologia , Proteínas do Citoesqueleto/metabolismo , Células HCT116 , Humanos , Leucemia Eritroblástica Aguda/patologia , Camundongos , Microtúbulos/metabolismo , Células NIH 3T3 , Análise de Sequência com Séries de Oligonucleotídeos , Regiões Promotoras Genéticas , Ativação TranscricionalRESUMO
Hepatitis C virus (HCV) encodes a polyprotein consisting of core, envelope (E1, E2, p7), and nonstructural polypeptides (NS2, NS3, NS4A, NS4B, NS5A, NS5B). The serine protease (NS3/NS4A), helicase (NS3), and polymerase (NS5B) constitute valid targets for antiviral therapy. We engineered BH3 interacting domain death agonist (BID), an apoptosis-inducing molecule, to contain a specific cleavage site recognized by the NS3/NS4A protease. Cleavage of the BID precursor molecule by the viral protease activated downstream apoptotic molecules of the mitochondrial pathway and triggered cell death. We extended this concept to cells transfected with an infectious HCV genome, hepatocytes containing HCV replicons, a Sindbis virus model for HCV, and finally HCV-infected mice with chimeric human livers. Infected mice injected with an adenovirus vector expressing modified BID exhibited HCV-dependent apoptosis in the human liver xenograft and considerable declines in serum HCV titers.