RESUMO
UMG1 is a unique epitope of CD43, not expressed by normal cells and tissues of haematopoietic and non-haematopoietic origin, except thymocytes and a minority (<5%) of peripheral blood T lymphocytes. By immunohistochemistry analysis of tissue microarray and pathology slides, we found high UMG1 expression in 20%-24% of diffuse large B-cell lymphomas (DLBCLs), including highly aggressive BCL2high and CD20low cases. UMG1 membrane expression was also found in DLBCL bone marrow-infiltrating cells and established cell lines. Targeting UMG1 with a novel asymmetric UMG1/CD3ε-bispecific T-cell engager (BTCE) induced redirected cytotoxicity against DLBCL cells and was synergistic with lenalidomide. We conclude that UMG1/CD3ε-BTCE is a promising therapeutic for DLBCLs.
Assuntos
Linfoma Difuso de Grandes Células B , Linfócitos T , Humanos , Linfócitos T/metabolismo , Linfoma Difuso de Grandes Células B/patologia , Imuno-HistoquímicaRESUMO
The identification of metazoan origins of DNA replication has so far been hampered by the lack of a suitable genetic screening and by the cumbersomeness of the currently available mapping procedures. Here we describe the construction of a library of nascent DNA, representative of all cellular origin sequences, and its utilization as a screening probe for origin identification in large genomic regions. The procedure developed was successfully applied to the human 5q31.1 locus, encoding for the IL-3 and GM-CSF genes. Two novel origins were identified and subsequently characterized by competitive PCR mapping, located approximately 3.5 kb downstream of the GM-CSF gene. The two origins (GM-CSF Ori1 and Ori2) were shown to interact with different members of the DNA prereplication complex. This observation reinforces the universal paradigm that initiation of DNA replication takes place at, or in close proximity to, the binding sites of the trans-acting initiator proteins.
Assuntos
Mapeamento Cromossômico/métodos , Replicação do DNA , DNA/genética , Biblioteca Gênica , Reação em Cadeia da Polimerase/métodos , Origem de Replicação , DNA/biossíntese , Proteínas de Ligação a DNA/análise , Proteínas de Ligação a DNA/genética , Evolução Molecular , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Humanos , Interleucina-3/genética , Hibridização de Ácido NucleicoRESUMO
In eukaryotes, DNA replication requires the regulated assembly of pre-replicative complexes (pre-RCs) onto DNA during G1 phase. Pre-RCs render the chromatin competent to replicate, yet it is only at the G1-S phase transition that protein-kinase complexes trigger the transition to DNA replication. Central to the formation of pre-RCs and regulation of DNA replication is the Cdc6 protein. Two recent studies have shown that Cdc6 is the long-sought factor that confers the competence to replicate in unfertilized Xenopus eggs.
Assuntos
Proteínas de Ciclo Celular/genética , Divisão Celular/genética , Replicação do DNA/genética , Células Eucarióticas/metabolismo , Proteínas de Saccharomyces cerevisiae , Animais , Proteínas de Ciclo Celular/metabolismo , Diferenciação Celular/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Substâncias Macromoleculares , Meiose/genéticaRESUMO
In yeasts, the replication protein Cdc6/Cdc18 is required for the initiation of DNA replication and also for coupling S phase with the following mitosis. In metazoans a role for Cdc6 has only been shown in S phase entry. Here we provide evidence that human Cdc6 (HuCdc6) also regulates the onset of mitosis, as overexpression of HuCdc6 in G(2) phase cells prevents entry into mitosis. This block is abolished when HuCdc6 is expressed together with a constitutively active Cyclin B/CDK1 complex or with Cdc25B or Cdc25C. An inhibitor of Chk1 kinase activity, UCN-01, overcomes the HuCdc6 mediated G(2) arrest indicating that HuCdc6 blocks cells in G(2) phase via a checkpoint pathway involving Chk1. When HuCdc6 is overexpressed in G(2), we detected phosphorylation of Chk1. Thus, HuCdc6 can trigger a checkpoint response, which could ensure that all DNA is replicated before mitotic entry. We also present evidence that the ability of HuCdc6 to block mitosis may be regulated by its phosphorylation.
Assuntos
Proteínas de Ciclo Celular/metabolismo , Replicação do DNA/fisiologia , Mitose/fisiologia , Proteínas Nucleares/metabolismo , Proteínas Quinases/metabolismo , Alcaloides/metabolismo , Proteínas de Ciclo Celular/genética , Quinase 1 do Ponto de Checagem , Dano ao DNA , Inibidores Enzimáticos/metabolismo , Células HeLa , Humanos , Microinjeções , Mutação , Proteínas Nucleares/genética , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Estaurosporina/análogos & derivadosRESUMO
A report on the British Society for Cell Biology (BSCB) meeting on 'Cell Biology and Neurobiology: A Meeting for Martin Raff', London, UK, 3-5 July 2002.
Assuntos
Biologia , Proteínas de Ligação a DNA , Neurobiologia , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos , Comportamento/fisiologia , Encéfalo/crescimento & desenvolvimento , Encéfalo/fisiologia , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Quinases Ciclina-Dependentes/antagonistas & inibidores , Inibidores Enzimáticos/uso terapêutico , Genes p53/genética , Genes p53/fisiologia , Humanos , Proteínas de Membrana/deficiência , Camundongos , Camundongos Knockout , Neurônios Motores/patologia , Neurônios Motores/fisiologia , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/fisiologia , Sistema Nervoso/enzimologia , Sistema Nervoso/crescimento & desenvolvimento , Malformações do Sistema Nervoso/diagnóstico , Malformações do Sistema Nervoso/genética , Neoplasias do Sistema Nervoso/diagnóstico , Neoplasias do Sistema Nervoso/tratamento farmacológico , Neoplasias do Sistema Nervoso/enzimologia , Neoplasias do Sistema Nervoso/genética , Fator de Transcrição 2 de Oligodendrócitos , Oligodendroglia/patologia , Oligodendroglia/fisiologia , Purinas/uso terapêutico , Roscovitina , Comportamento Social , Canais de Cátion TRPCRESUMO
In eukaryotes, the initiation of DNA replication involves the ordered assembly on chromatin of pre-replicative complexes (pre-RCs), including the origin recognition complex (ORC), Cdc6, Cdt1 and the minichromosome maintenance proteins (MCMs). In light of its indispensable role in the formation of pre-RCs, Cdc6 binding to chromatin represents a key step in the regulation of DNA replication and cell proliferation. Here, we study the human Cdc6 (HuCdc6) protein during programmed cell death (apoptosis). We find that HuCdc6, but not HuOrc2 (a member of the ORC) or HuMcm5 (one of the MCMs), is specifically cleaved in several human cell lines induced to undergo apoptosis by a variety of stimuli. Expression of caspase-uncleavable mutant HuCdc6 attenuates apoptosis, delaying cell death. Therefore, an important function for cleavage of HuCdc6 is to prevent a wounded cell from replicating and to facilitate death.
