RESUMEN
A genomic locus 8 kb downstream of the transcription factor GFI1B (Growth Factor Independence 1B) predisposes to clonal hematopoiesis and myeloproliferative neoplasms. One of the most significantly associated polymorphisms in this region is rs621940-G. GFI1B auto-represses GFI1B, and altered GFI1B expression contributes to myeloid neoplasms. We studied whether rs621940-G affects GFI1B expression and growth of immature cells. GFI1B ChIP-seq showed clear binding to the rs621940 locus. Preferential binding of various hematopoietic transcription factors to either the rs621940-C or -G allele was observed, but GFI1B showed no preference. In gene reporter assays the rs621940 region inhibited GFI1B promoter activity with the G-allele having less suppressive effects compared to the C-allele. However, CRISPR-Cas9 mediated deletion of the locus in K562 cells did not alter GFI1B expression nor auto-repression. In healthy peripheral blood mononuclear cells GFI1B expression did not differ consistently between the rs621940 alleles. Long range and targeted deep sequencing did not detect consistent effects of rs621940-G on allelic GFI1B expression either. Finally, we observed that myeloid colony formation was not significantly affected by either rs621940 allele in 193 healthy donors. Together, these findings show no evidence that rs621940 or its locus affect GFI1B expression, auto-repression or growth of immature myeloid cells.
Asunto(s)
Predisposición Genética a la Enfermedad , Trastornos Mieloproliferativos/genética , Neoplasias/genética , Proteínas Proto-Oncogénicas/genética , Proteínas Represoras/genética , Adulto , Anciano , Alelos , Sistemas CRISPR-Cas/genética , Femenino , Regulación de la Expresión Génica/genética , Genoma Humano/genética , Genotipo , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Células K562 , Masculino , Persona de Mediana Edad , Células Mieloides/metabolismo , Células Mieloides/patología , Trastornos Mieloproliferativos/patología , Neoplasias/patología , Fagocitosis/genética , Polimorfismo de Nucleótido Simple/genética , Adulto JovenRESUMEN
We assessed the prognostic impact of TET2 mutations and mRNA expression in a prospective cohort of 357 adult AML patients < 60 years of age enrolled in the European Organization For Research and Treatment of Cancer (EORTC)/Gruppo Italiano Malattie Ematologiche dell' Adulto (GIMEMA) AML-12 06991 clinical trial. In addition the co-occurrence with other genetic defects and the functional consequences of TET2 mutations were investigated. TET2 mutations occurred in 7.6 % of the patients and were an independent marker of poor prognosis (p = 0.024). TET2 and IDH1/2 mutations strongly associated with aberrations in the DNA methyltransferase DNMT3A. Functional studies confirmed previous work that neither nonsense truncations, nor missense TET2 mutations, induced 5-hydroxymethylcytosine formation. In addition, we now show that mutant TET2 forms did not act in a dominant negative manner when co-expressed with the wild-type protein. Finally, as loss-of-function TET2 mutations predicted poor outcome, we questioned whether low TET2 mRNA expression in cases of AML without TET2 mutations would affect overall survival. Notably, also AML patients with low TET2 mRNA expression levels showed inferior overall survival.
Asunto(s)
Proteínas de Unión al ADN/genética , Regulación Leucémica de la Expresión Génica , Leucemia Mieloide Aguda/genética , Mutación , Proteínas de Neoplasias/genética , Proteínas Proto-Oncogénicas/genética , 5-Metilcitosina/análogos & derivados , Adolescente , Adulto , Animales , Células COS , Chlorocebus aethiops , Ensayos Clínicos como Asunto , Citosina/análogos & derivados , Citosina/análisis , ADN (Citosina-5-)-Metiltransferasas/genética , ADN Metiltransferasa 3A , Proteínas de Unión al ADN/biosíntesis , Proteínas de Unión al ADN/fisiología , Dioxigenasas , Femenino , Humanos , Isocitrato Deshidrogenasa/genética , Estimación de Kaplan-Meier , Leucemia Mieloide Aguda/tratamiento farmacológico , Leucemia Mieloide Aguda/mortalidad , Masculino , Estudios Multicéntricos como Asunto , Proteínas de Neoplasias/biosíntesis , Proteínas de Neoplasias/fisiología , Pronóstico , Estudios Prospectivos , Proteínas Proto-Oncogénicas/biosíntesis , Proteínas Proto-Oncogénicas/fisiología , ARN Mensajero/biosíntesis , ARN Neoplásico/biosíntesis , Proteínas Recombinantes de Fusión/metabolismo , Transfección , Adulto JovenAsunto(s)
Proteínas Portadoras/genética , Proteínas de Unión al ADN/genética , Variación Genética , Neoplasias Hematológicas/genética , Proteínas Nucleares/genética , Proteínas Represoras/genética , Factores de Transcripción/genética , Estudios de Cohortes , Proteína Potenciadora del Homólogo Zeste 2 , Frecuencia de los Genes , Predisposición Genética a la Enfermedad , Variación Genética/fisiología , Humanos , Síndromes Mielodisplásicos/genética , Trastornos Mieloproliferativos/genética , Proteínas de Neoplasias , Complejo Represivo Polycomb 2 , Proteínas del Grupo Polycomb , Polimorfismo de Nucleótido SimpleRESUMEN
Myelodysplastic syndromes (MDS) represent a heterogeneous group of neoplastic hematopoietic disorders. Several recurrent chromosomal aberrations have been associated with MDS, but the genes affected have remained largely unknown. To identify relevant genetic lesions involved in the pathogenesis of MDS, we conducted SNP array-based genomic profiling and genomic sequencing in 102 individuals with MDS and identified acquired deletions and missense and nonsense mutations in the TET2 gene in 26% of these individuals. Using allele-specific assays, we detected TET2 mutations in most of the bone marrow cells (median 96%). In addition, the mutations were encountered in various lineages of differentiation including CD34(+) progenitor cells, suggesting that TET2 mutations occur early during disease evolution. In healthy tissues, TET2 expression was shown to be elevated in hematopoietic cells with highest expression in granulocytes, in line with a function in myelopoiesis. We conclude that TET2 is the most frequently mutated gene in MDS known so far.
