RESUMO
The Philadelphia chromosome (Ph) encoding the oncogenic BCR-ABL1 kinase defines a subset of acute lymphoblastic leukemia (ALL) with a particularly unfavorable prognosis. ALL cells are derived from B cell precursors in most cases and typically carry rearranged immunoglobulin heavy chain (IGH) variable (V) region genes devoid of somatic mutations. Somatic hypermutation is restricted to mature germinal center B cells and depends on activation-induced cytidine deaminase (AID). Studying AID expression in 108 cases of ALL, we detected AID mRNA in 24 of 28 Ph(+) ALLs as compared with 6 of 80 Ph(-) ALLs. Forced expression of BCR-ABL1 in Ph(-) ALL cells and inhibition of the BCR-ABL1 kinase showed that aberrant expression of AID depends on BCR-ABL1 kinase activity. Consistent with aberrant AID expression in Ph(+) ALL, IGH V region genes and BCL6 were mutated in many Ph(+) but unmutated in most Ph(-) cases. In addition, AID introduced DNA single-strand breaks within the tumor suppressor gene CDKN2B in Ph(+) ALL cells, which was sensitive to BCR-ABL1 kinase inhibition and silencing of AID expression by RNA interference. These findings identify AID as a BCR-ABL1-induced mutator in Ph(+) ALL cells, which may be relevant with respect to the particularly unfavorable prognosis of this leukemia subset.
Assuntos
Linfócitos B/imunologia , Citidina Desaminase/metabolismo , Regulação Neoplásica da Expressão Gênica/imunologia , Cromossomo Filadélfia , Leucemia-Linfoma Linfoblástico de Células Precursoras/imunologia , Proteínas Tirosina Quinases/genética , Linfócitos B/metabolismo , Sequência de Bases , Western Blotting , Análise Mutacional de DNA , Proteínas de Ligação a DNA/genética , Citometria de Fluxo , Proteínas de Fusão bcr-abl , Genes myc/genética , Humanos , Região Variável de Imunoglobulina/genética , Dados de Sequência Molecular , Mutação/genética , Análise de Sequência com Séries de Oligonucleotídeos , Oligonucleotídeos , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismo , Proteínas Tirosina Quinases/metabolismo , Proteínas Proto-Oncogênicas c-bcl-6 , Interferência de RNA , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Alinhamento de SequênciaRESUMO
A number of recent studies identified nuclear factors that together have the unique ability to induce pluripotency in differentiated cell types. However, little is known about the factors that are needed to maintain human embryonic stem (ES) cells in an undifferentiated state. In a search for such requirements, we performed a comprehensive meta-analysis of publicly available SAGE and microarray data. The rationale for this analysis was to identify genes that are exclusively expressed in human ES cell lines compared to 30 differentiated tissue types. The WNT receptor FZD7 was found among the genes with an ES cell-specific expression profile in both SAGE and microarray analyses. Subsequent validation by quantitative RT-PCR and flow cytometry confirmed that FZD7 mRNA levels in human ES cells are up to 200-fold higher compared to differentiated cell types. ShRNA-mediated knockdown of FZD7 in human ES cells induced dramatic changes in the morphology of ES cell colonies, perturbation of expression levels of germ layer-specific marker genes, and a rapid loss of expression of the ES cell-specific transcription factor OCT4. These findings identify the WNT receptor FZD7 as a novel ES cell-specific surface antigen with a likely important role in the maintenance of ES cell self-renewal capacity.