RESUMO
The t(11;14)(q13;q32) between the BCL-1 and immunoglobulin heavy chain gene (IgH) loci in mantle cell lymphoma (MCL) are believed to be mediated by the mechanism of V(D)J recombination similar to the t(14; 18) in follicular lymphoma (FL). We have recently shown that the t(14;18) event creates staggered double-strand breaks in the BCL-2 locus, and that the t(14;18) junctions contain templated nucleotide insertions (T-nucleotides; U. Jäger et al., Blood, 95: 3520-3529, 2000). Reasoning that the earlier (pregerminal center) B-cell origin of MCL might be reflected in a different molecular structure of the chromosomal breakpoints, we PCR-amplified diagnostic samples from 93 patients. Thirty-six samples (39%) were positive for the direct (BCL-1/J(H)) and 23 for both direct and reciprocal (D(H)/BCL-1) junctions. The breaks on chromosome 14 exhibited features of V(D)J-mediated recombination as shown by D(H) and J(H) coding end processing. However, duplications of BCL-1 sequences in 39% of the 23 patients indicate staggered double-strand breaks in the major translocation cluster region (MTC). This is incompatible with V(D)J recombination and indicates a different mechanism of cleavage. The use of J(H)6 in the junctions (39%) was similar to that in the immunoglobulin genes of normal B cells and B-CLL, but considerably less than in FL. Only 2 of 36 samples contained a BCL-1/DJ(H) rearrangement, which was indicative of a previous DJ(H) rearrangement. Most importantly, 19% of the BCL-1/IgH junctions with inserts of > or =5 nucleotides contained error-prone copies (T-nucleotides) of 8-12 nucleotides originating from the surrounding BCL-1 or IgH regions, a lower rate than in FL. No correlation was found between the addition of T-nucleotides and the rate of somatic mutation in the immunoglobulin genes. We conclude that the t(11;14) and t(14;18) use the same basic mechanism of translocation including V(D)J-mediated recombination, double-strand staggered breaks, and template-dependent, error-prone DNA-synthesis. However, the distinct differences in the utilization of J(H) regions suggest that the t(11;14) occurs predominantly during an attempted primary D(H)-J(H) rearrangement in early B cells, whereas the t(14;18) mostly occurs during secondary rearrangement. This is in agreement with the pregerminal center B-cell origin of MCL.
Assuntos
Genes de Imunoglobulinas/genética , Cadeias Pesadas de Imunoglobulinas/genética , Cadeias J de Imunoglobulina/genética , Linfoma de Célula do Manto/genética , Translocação Genética/genética , Sequência de Bases , Quebra Cromossômica/genética , Cromossomos Humanos Par 11 , Cromossomos Humanos Par 14 , Elementos de DNA Transponíveis/genética , Rearranjo Gênico de Cadeia Pesada de Linfócito B/genética , Genes bcl-1/genética , Humanos , Idiótipos de Imunoglobulinas/genética , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Moldes GenéticosRESUMO
The Raf-1 kinase plays a key role in relaying proliferation signals elicited by mitogens or oncogenes. Raf-1 is regulated by complex and incompletely understood mechanisms including phosphorylation. A number of studies have indicated that phosphorylation of serines 259 and 621 can inhibit the Raf-1 kinase. We show that both serines are hypophosphorylated during early mitogenic stimulation and that hypophosphorylation correlates with peak Raf-1 activation. Concentrations of okadaic acid that selectively inhibit protein phosphatase 2A (PP2A) induce phosphorylation of these residues and prevent maximal activation of the Raf-1 kinase. This effect is mediated via phosphorylation of serine 259. The PP2A core heterodimer forms complexes with Raf-1 in vivo and in vitro. These data identify PP2A as a positive regulator of Raf-1 activation and are the first indication that PP2A may support the activation of an associated kinase.
