RESUMEN
SATB1 (Special A-T rich Binding protein 1) is a cell type-specific factor that regulates the genetic network in developing T cells and neurons. In T cells, SATB1 is required for lineage commitment, VDJ recombination, development and maturation. Considering that its expression varies during B-cell differentiation, the involvement of SATB1 needs to be clarified in this lineage. Using a KO mouse model in which SATB1 was deleted from the pro-B-cell stage, we examined the consequences of SATB1 deletion in naive and activated B-cell subsets. Our model indicates first, unlike its essential function in T cells, that SATB1 is dispensable for B-cell development and the establishment of a broad IgH repertoire. Second, we show that SATB1 exhibits an ambivalent function in mature B cells, acting sequentially as a positive and negative regulator of Ig gene transcription in naive and activated cells, respectively. Third, our study indicates that the negative regulatory function of SATB1 in B cells extends to the germinal center response, in which this factor limits somatic hypermutation of Ig genes.
Asunto(s)
Proteínas de Unión a la Región de Fijación a la Matriz , Animales , Ratones , Proteínas de Unión a la Región de Fijación a la Matriz/genética , Proteínas de Unión a la Región de Fijación a la Matriz/metabolismo , Redes Reguladoras de Genes , Linfocitos T/metabolismo , Factores de Transcripción/metabolismo , Cromatina/metabolismoAsunto(s)
Linfocitos B/fisiología , Elementos de Facilitación Genéticos/genética , Cadenas Pesadas de Inmunoglobulina/genética , Cadenas kappa de Inmunoglobulina/genética , Células Precursoras de Linfocitos B/fisiología , Secuencias Reguladoras de Ácidos Nucleicos/genética , Recombinación V(D)J/genética , Animales , Diferenciación Celular , Silenciador del Gen , Genes RAG-1/genética , Cambio de Clase de Inmunoglobulina , Ratones , Ratones Noqueados , Activación TranscripcionalAsunto(s)
Región de Flanqueo 3'/genética , Región de Flanqueo 5'/genética , Linfocitos B/inmunología , Elementos de Facilitación Genéticos , Cadenas Pesadas de Inmunoglobulina/genética , Región de Control de Posición/genética , Secuencias Reguladoras de Ácidos Nucleicos/genética , Animales , Células Cultivadas , Femenino , Proteínas de Homeodominio/fisiología , Cambio de Clase de Inmunoglobulina/genética , Masculino , Ratones , Ratones Noqueados , Eliminación de Secuencia , Hipermutación Somática de Inmunoglobulina , Transcripción GenéticaRESUMEN
In B-lineage cells, the cytidine deaminase AID not only generates somatic mutations to variable regions of Ig genes but also inflicts, at a lower frequency, mutations to several non-Ig genes named AID off-targets, which include proto-oncogenes. High-throughput sequencing should be in principle the method of choice to detect and document these rare nucleotide substitutions. So far, high-throughput sequencing-based methods are impaired by a global sequencing error rate that usually covers the real mutation rate of AID off-target genes in activated B cells. We demonstrate the validity of a per-base background subtraction method called detection of minor variants by error correction (DeMinEr), which uses deep sequencing data from mutated and nonmutated samples to correct the substitution frequency at each nucleotide position along the sequenced region. Our DeMinEr method identifies somatic mutations at a frequency down to 0.02% at any nucleotide position within two off-target genes: Cd83 and Bcl6 Biological models and control conditions such as AID- and UNG-deficient mice validate the specificity and the sensitivity of our method. The high resolution and robustness of DeMinEr enable us to document fine effects such as age-dependent accumulation of mutations in these oncogenes in the mouse.