RESUMO
FACT, in addition to its role in transcription, is likely implicated in both transcription-coupled nucleotide excision repair and DNA double strand break repair. Here, we present evidence that FACT could be directly involved in Base Excision Repair and elucidate the chromatin remodeling mechanisms of FACT during BER. We found that, upon oxidative stress, FACT is released from transcription related protein complexes to get associated with repair proteins and chromatin remodelers from the SWI/SNF family. We also showed the rapid recruitment of FACT to the site of damage, coincident with the glycosylase OGG1, upon the local generation of oxidized DNA. Interestingly, FACT facilitates uracil-DNA glycosylase in the removal of uracil from nucleosomal DNA thanks to an enhancement in the remodeling activity of RSC. This discloses a novel property of FACT wherein it has a co-remodeling activity and strongly enhances the remodeling capacity of the chromatin remodelers. Altogether, our data suggest that FACT may acts in concert with RSC to facilitate excision of DNA lesions during the initial step of BER.
Assuntos
Reparo do DNA/genética , Proteínas de Ligação a DNA/genética , Proteínas de Grupo de Alta Mobilidade/genética , Histonas/genética , Transcrição Gênica , Fatores de Elongação da Transcrição/genética , Animais , Cromatina/genética , Montagem e Desmontagem da Cromatina/genética , Proteínas Cromossômicas não Histona/genética , Dano ao DNA/genética , Proteínas de Ligação a DNA/biossíntese , Células HeLa , Proteínas de Grupo de Alta Mobilidade/biossíntese , Histonas/metabolismo , Humanos , Nucleossomos/genética , Estresse Oxidativo/genética , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Fatores de Transcrição/genética , Fatores de Elongação da Transcrição/biossíntese , Uracila/metabolismo , Xenopus laevisRESUMO
BACKGROUND AND OBJECTIVES: To clarify the relationship between splenic (SMZL) and nodal marginal zone (NMZL) lymphomas, we analyzed immunoglobulin variable heavy chain (VH) gene usage and mutation patterns in these tumors. DESIGN AND METHODS: VH genes were cloned and sequenced from 49 lymphoma samples (35 SMZL and 14 NMZL). RESULTS: A biased usage of VH gene was found with overrepresentation of VH1 in SMZL cases (13/35) and VH4 in NMZL cases (7/14). Evidence for antigen driven mutations was identified in 8 SMZL and 4 NMZL cases. Three cases out of 18 with clones analyzed from spleen and peripheral blood demonstrated intra-clonal diversity, with evidence of clonal selection in one case, indicating the possibility of antigen-driven clonal expansion. Eleven SMZL cases (31%) but only 2 NMZL (14%) cases were unmutated. No differences in clinical outcome and overall survival were found between the unmutated and mutated cases. INTERPRETATION AND CONCLUSIONS: The pattern of somatic mutation and the VH gene segment usage appear to differ between SMZL and NMZL, suggesting that these are distinct pathological entities. Moreover, a biased usage of certain sequences suggests that tumor cells in SMZL may be subjected to antigen selection.
Assuntos
Cadeias Pesadas de Imunoglobulinas/genética , Região Variável de Imunoglobulina/genética , Doenças Linfáticas/genética , Linfoma de Células B/genética , Neoplasias Esplênicas/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Anergia Clonal/genética , Feminino , Genes de Imunoglobulinas/fisiologia , Humanos , Doenças Linfáticas/sangue , Linfoma de Células B/sangue , Masculino , Pessoa de Meia-Idade , Neoplasias Esplênicas/sangueRESUMO
The human corticotropin-releasing factor receptor type 1 (hCRF-R1) functional transcript is mainly expressed in the anterior pituitary corticotrophs, a tissue usually not available for clinical investigation. Splice variants translated into defective isoforms of the receptor have been described in few peripheral tissues. The aim of this work was to determine whether peripheral white blood cells from healthy individuals, an accessible tissue for clinical investigation, were suitable for the analysis of the hCRF-R1 transcript and gene. We report that: (i) specific amplification of the hCRF-R1 transcript from peripheral white blood cells mRNAs is feasible; (ii) this transcript is similar to the functional transcript; (iii) the draft sequence of chromosome 17 and unrelated sequences allow direct sequencing of all 14 exons of the gene, adjacent splice sites and related branch points. In conclusion, these approaches would be suitable for studies in patients having isolated secondary glucocorticoids deficiency to implicate the hCRH-R1 in the etiology of the disease.
Assuntos
DNA/isolamento & purificação , Leucócitos , RNA Mensageiro/isolamento & purificação , Receptores de Hormônio Liberador da Corticotropina/genética , Primers do DNA , Humanos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodosRESUMO
In this study, we explored the telomeric changes that occur in B-chronic lymphocytic leukemia (B-CLL), in which telomere length has recently been demonstrated to be a powerful prognostic marker. We carried out a transcriptomic analysis of telomerase components (hTERT and DYSKERIN), shelterin proteins (TRF1, TRF2, hRAP1, TIN2, POT1, and TPP1), and a set of multifunctional proteins involved in telomere maintenance (hEST1A, MRE11, RAD50, Ku80, and RPA1) in peripheral B cells from 42 B-CLL patients and 20 healthy donors. We found that, in B-CLL cells, the expressions of hTERT, DYSKERIN, TRF1, hRAP1, POT1, hEST1A, MRE11, RAD50, and KU80 were more than 2-fold reduced (P < .001), contrasting with the higher expression of TPP1 and RPA1 (P < .001). This differential expression pattern suggests that both telomerase down-regulation and changes in telomeric proteins composition are involved in the pathogenesis of B-CLL.