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J Exp Med ; 216(5): 1050-1060, 2019 May 06.
Artigo em Inglês | MEDLINE | ID: mdl-30914438


Studies of allelic variation underlying genetic blood disorders have provided important insights into human hematopoiesis. Most often, the identified pathogenic mutations result in loss-of-function or missense changes. However, assessing the pathogenicity of noncoding variants can be challenging. Here, we characterize two unrelated patients with a distinct presentation of dyserythropoietic anemia and other impairments in hematopoiesis associated with an intronic mutation in GATA1 that is 24 nucleotides upstream of the canonical splice acceptor site. Functional studies demonstrate that this single-nucleotide alteration leads to reduced canonical splicing and increased use of an alternative splice acceptor site that causes a partial intron retention event. The resultant altered GATA1 contains a five-amino acid insertion at the C-terminus of the C-terminal zinc finger and has no observable activity. Collectively, our results demonstrate how altered splicing of GATA1, which reduces levels of the normal form of this master transcription factor, can result in distinct changes in human hematopoiesis.

Elife ; 72018 11 23.
Artigo em Inglês | MEDLINE | ID: mdl-30468428


DNA methylation plays an essential role in mammalian genomes and expression of the responsible enzymes is tightly controlled. Deregulation of the de novo DNA methyltransferase DNMT3B is frequently observed across cancer types, yet little is known about its ectopic genomic targets. Here, we used an inducible transgenic mouse model to delineate rules for abnormal DNMT3B targeting, as well as the constraints of its activity across different cell types. Our results explain the preferential susceptibility of certain CpG islands to aberrant methylation and point to transcriptional state and the associated chromatin landscape as the strongest predictors. Although DNA methylation and H3K27me3 are usually non-overlapping at CpG islands, H3K27me3 can transiently co-occur with DNMT3B-induced DNA methylation. Our genome-wide data combined with ultra-deep locus-specific bisulfite sequencing suggest a distributive activity of ectopically expressed Dnmt3b that leads to discordant CpG island hypermethylation and provides new insights for interpreting the cancer methylome.

Am J Hematol ; 93(11): 1358-1367, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30117174


The B/T subtype of mixed phenotype acute leukemia (B/T MPAL) is defined by co-expression of antigens of both B- and T-cell lineages on leukemic blasts. Although it has been suggested that multilineage antigen expression portends poor response to chemotherapy, the clinical characteristics and driver mutations that underlie the pathogenesis of this rare subtype of acute leukemia are scarcely known. We identified nine cases of B/T MPAL from multiple institutions and correlated clinical and immunophenotypic findings with next-generation sequencing data. We report that B/T MPAL commonly presents with lymphadenopathy in adolescence and young adulthood. While the tumors have diverse cytogenetic and genomic perturbations, recurrent acquired aberrations include mutations in the putative transcriptional regulator PHF6 and the JAK-STAT and Ras signaling pathways. Alterations were also identified in genes encoding hematopoietic transcription factors, cell cycle regulators/tumor suppressors, and chromatin modifying enzymes. The genomic landscape of B/T MPAL strongly resembles that of T-ALL subgroups associated with early developmental arrest, while genetic alterations that are common in B-ALL were rarely seen. Two-thirds of the patients responded to ALL-based chemotherapy with or without stem cell transplantation. Our observations lay the groundwork for further study of the unique biology and clinical trajectory of B/T MPAL.

Wei Sheng Yan Jiu ; 35(6): 798-801, 2006 Nov.
Artigo em Chinês | MEDLINE | ID: mdl-17290771


OBJECTIVE: In order to provide a rapid and selectivity method for the determination of clenbuterol(CBL), an indirect competitive time-resolved fluoroimmunoassay (TRFIA) was developed. METHODS: Anti-CBL antibody, was raised by immunization against CBL-BSA in rabbits. CBL-OVA was coated by physical adsorption onto the microtitre plate, CBL or sample with CBL as a competitor. Both them were incubated with limited anti- CBL antibody. and a goat antirabbit IgG-Eu3+ conjugate was used as a tracer. RESULTS: The sensitivity of CBL-TRFIA was 0.01microg/L, and the recovery rate was 99.7%. RSD of CBL-TRFIA was 3.9% . The sensitivity of CBL-TRFIA provided a linear response from 0.01 - 25microg/L, with ED50 of (1.47+/-0.11) microg/L or ED80 of (0.07+/-0.01)microg/L and ED, of (23.6+/- 0.56) microg/L. The cross reactivity of the CBL-TRFIA with salbutamol, epinephrine hydrochloride and epinephrine bitartrate was negligible, while that with isoprenaline hydrochloride was 0.01% . Both CBL-TRFIA and CBL-ELISA test were applied for the quantitative measurement of CBL in the same urine, and the coefficient of correlation was 0.932. CONCLUSION: The CBL-TRFIA could be applied to detect the CBL in urine and it is useful to screening easily for CBL contamination in meat or foods.

Clembuterol/urina , Fluorimunoensaio/métodos , Animais , Masculino , Coelhos , Sensibilidade e Especificidade