RESUMO
Somatic cell nuclear transfer and transcription-factor-based reprogramming revert adult cells to an embryonic state, and yield pluripotent stem cells that can generate all tissues. Through different mechanisms and kinetics, these two reprogramming methods reset genomic methylation, an epigenetic modification of DNA that influences gene expression, leading us to hypothesize that the resulting pluripotent stem cells might have different properties. Here we observe that low-passage induced pluripotent stem cells (iPSCs) derived by factor-based reprogramming of adult murine tissues harbour residual DNA methylation signatures characteristic of their somatic tissue of origin, which favours their differentiation along lineages related to the donor cell, while restricting alternative cell fates. Such an 'epigenetic memory' of the donor tissue could be reset by differentiation and serial reprogramming, or by treatment of iPSCs with chromatin-modifying drugs. In contrast, the differentiation and methylation of nuclear-transfer-derived pluripotent stem cells were more similar to classical embryonic stem cells than were iPSCs. Our data indicate that nuclear transfer is more effective at establishing the ground state of pluripotency than factor-based reprogramming, which can leave an epigenetic memory of the tissue of origin that may influence efforts at directed differentiation for applications in disease modelling or treatment.
Assuntos
Epigênese Genética , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Animais , Diferenciação Celular/genética , Linhagem da Célula/genética , Reprogramação Celular/genética , Metilação de DNA/genética , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Genoma/genética , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Técnicas de Transferência Nuclear , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
To identify new immortalizing genes with potential roles in tumorigenesis, we performed a genetic screen aimed to bypass the rapid and tight senescence arrest of primary fibroblasts deficient for the oncogene Bmi1. We identified the T-box member TBX2 as a potent immortalizing gene that acts by downregulating Cdkn2a (p19(ARF)). TBX2 represses the Cdkn2a (p19(ARF)) promoter and attenuates E2F1, Myc or HRAS-mediated induction of Cdkn2a (p19(ARF)). We found TBX2 to be amplified in a subset of primary human breast cancers, indicating that it might contribute to breast cancer development.
Assuntos
Adenocarcinoma/genética , Neoplasias da Mama/genética , Proteínas de Ciclo Celular/fisiologia , Senescência Celular/genética , Cromossomos Humanos Par 17/genética , Proteínas de Ligação a DNA , Amplificação de Genes , Regulação Neoplásica da Expressão Gênica , Proteínas de Neoplasias/fisiologia , Biossíntese de Proteínas , Proteínas Repressoras/fisiologia , Proteínas com Domínio T/fisiologia , Adenocarcinoma/metabolismo , Animais , Neoplasias da Mama/metabolismo , Células COS , Proteínas de Transporte/genética , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/isolamento & purificação , Transformação Celular Neoplásica/genética , Células Cultivadas , Inibidor p16 de Quinase Dependente de Ciclina , Fatores de Transcrição E2F , Fator de Transcrição E2F1 , Feminino , Fibroblastos/citologia , Fibroblastos/metabolismo , Deleção de Genes , Genes BRCA1 , Humanos , Camundongos , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/isolamento & purificação , Síndromes Neoplásicas Hereditárias/genética , Proteínas Nucleares/genética , Oncogenes , Complexo Repressor Polycomb 1 , Regiões Promotoras Genéticas , Proteínas/genética , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-myc/antagonistas & inibidores , Proteínas Proto-Oncogênicas p21(ras)/antagonistas & inibidores , Proteínas Repressoras/genética , Proteínas Repressoras/isolamento & purificação , Proteína 1 de Ligação ao Retinoblastoma , Proteínas com Domínio T/genética , Proteínas com Domínio T/isolamento & purificação , Fator de Transcrição DP1 , Fatores de Transcrição/antagonistas & inibidores , Transfecção , Células Tumorais Cultivadas , Proteína Supressora de Tumor p14ARFRESUMO
A major goal in human genetics is to understand the role of common genetic variants in susceptibility to common diseases. This will require characterizing the nature of gene variation in human populations, assembling an extensive catalogue of single-nucleotide polymorphisms (SNPs) in candidate genes and performing association studies for particular diseases. At present, our knowledge of human gene variation remains rudimentary. Here we describe a systematic survey of SNPs in the coding regions of human genes. We identified SNPs in 106 genes relevant to cardiovascular disease, endocrinology and neuropsychiatry by screening an average of 114 independent alleles using 2 independent screening methods. To ensure high accuracy, all reported SNPs were confirmed by DNA sequencing. We identified 560 SNPs, including 392 coding-region SNPs (cSNPs) divided roughly equally between those causing synonymous and non-synonymous changes. We observed different rates of polymorphism among classes of sites within genes (non-coding, degenerate and non-degenerate) as well as between genes. The cSNPs most likely to influence disease, those that alter the amino acid sequence of the encoded protein, are found at a lower rate and with lower allele frequencies than silent substitutions. This likely reflects selection acting against deleterious alleles during human evolution. The lower allele frequency of missense cSNPs has implications for the compilation of a comprehensive catalogue, as well as for the subsequent application to disease association.
Assuntos
Polimorfismo Genético , Alelos , Evolução Biológica , Frequência do Gene , Genes , Variação Genética , Humanos , Proteínas/genética , Análise de Sequência de DNARESUMO
The product of the Philadelphia chromosome (Ph) translocation, the BCR/ABL oncogene, exists in three principal forms (P190, P210, and P230 BCR/ABL) that are found in distinct forms of Ph-positive leukemia, suggesting the three proteins have different leukemogenic activity. We have directly compared the tyrosine kinase activity, in vitro transformation properties, and in vivo leukemogenic activity of the P190, P210, and P230 forms of BCR/ABL. P230 exhibited lower intrinsic tyrosine kinase activity than P210 and P190. Although all three oncogenes transformed both myeloid (32D cl3) and lymphoid (Ba/F3) interleukin (IL)-3-dependent cell lines to become independent of IL-3 for survival and growth, their ability to stimulate proliferation of Ba/F3 lymphoid cells differed and correlated directly with tyrosine kinase activity. In a murine bone marrow transduction/transplantation model, the three forms of BCR/ABL were equally potent in the induction of a chronic myeloid leukemia (CML)-like myeloproliferative syndrome in recipient mice when 5-fluorouracil (5-FU)-treated donors were used. Analysis of proviral integration showed the CML-like disease to be polyclonal and to involve multiple myeloid and B lymphoid lineages, implicating a primitive multipotential target cell. Secondary transplantation revealed that only certain minor clones gave rise to day 12 spleen colonies and induced disease in secondary recipients, suggesting heterogeneity among the target cell population. In contrast, when marrow from non- 5-FU-treated donors was used, a mixture of CML-like disease, B lymphoid acute leukemia, and macrophage tumors was observed in recipients. P190 BCR/ABL induced lymphoid leukemia with shorter latency than P210 or P230. The lymphoid leukemias and macrophage tumors had provirus integration patterns that were oligo- or monoclonal and limited to the tumor cells, suggesting a lineage-restricted target cell with a requirement for additional events in addition to BCR/ABL transduction for full malignant transformation. These results do not support the hypothesis that P230 BCR/ABL induces a distinct and less aggressive form of CML in humans, and suggest that the rarity of P190 BCR/ABL in human CML may reflect infrequent BCR intron 1 breakpoints during the genesis of the Ph chromosome in stem cells, rather than intrinsic differences in myeloid leukemogenicity between P190 and P210.
Assuntos
Proteínas de Fusão bcr-abl/fisiologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/etiologia , Animais , Divisão Celular , Linhagem Celular Transformada , Modelos Animais de Doenças , Proteínas de Fusão bcr-abl/genética , Proteínas de Fusão bcr-abl/metabolismo , Humanos , Leucemia Linfoide/fisiopatologia , Linfócitos/citologia , Camundongos , Camundongos Endogâmicos BALB C , Oncogenes , Isoformas de Proteínas , Proteínas Tirosina Quinases/metabolismo , SíndromeRESUMO
In tumor cells from virtually all patients with chronic myelogenous leukemia, the Philadelphia chromosome, a fusion of chromosomes 9 and 22, directs the synthesis of the P210bcr/abl protein. The protein-tyrosine kinase activity and hybrid structure of P210bcr/abl are similar to the oncogene product of the Abelson murine leukemia virus, P160gag/v-abl, which induces acute lymphomas. To determine whether P210bcr/abl can induce chronic myelogenous leukemia, murine bone marrow was infected with a retrovirus encoding P210bcr/abl and transplanted into irradiated syngeneic recipients. Transplant recipients developed several hematologic malignancies; prominent among them was a myeloproliferative syndrome closely resembling the chronic phase of human chronic myelogenous leukemia. Tumor tissue from diseased mice harbored the provirus encoding P210bcr/abl. These results demonstrate that P210bcr/abl expression can induce chronic myelogenous leukemia. Retrovirus-mediated expression of the protein provides a murine model system for further analysis of the disease.
Assuntos
Proteínas de Fusão bcr-abl/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Cromossomo Filadélfia , Animais , Medula Óssea/microbiologia , Medula Óssea/patologia , Linhagem Celular , DNA de Neoplasias/isolamento & purificação , DNA Viral/isolamento & purificação , Humanos , Leucemia Experimental/genética , Leucemia Experimental/patologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Camundongos , Transtornos Mieloproliferativos/microbiologia , Transtornos Mieloproliferativos/patologia , Retroviridae/genética , Baço/microbiologia , Transdução GenéticaRESUMO
The v-abl oncogene of the Abelson murine leukemia virus (A-MuLV) is known to efficiently transform NIH/3T3 fibroblasts in vitro and to cause an acute lymphosarcoma in susceptible murine hosts. The role of its relative, the bcr/abl gene product, in the etiology of human chronic myelogenous leukemia (CML) remains speculative. To assess the transforming properties of the bcr/abl gene product, complementary DNA clones encoding the CML-specific P210 bcr/abl protein were expressed in NIH/3T3 fibroblasts. In contrast to the v-abl oncogene product P160, the P210 bcr/abl gene product did not transform NIH/3T3 cells. Cell lines were isolated that expressed high levels of the P210 bcr/abl protein but were morphologically normal. During the course of these experiments, a transforming recombinant of bcr/abl was isolated which fuses gag determinants derived from helper virus to the NH2-terminus of the bcr/abl protein. This suggests that a property of viral gag sequences, probably myristylation-dependent membrane localization, must be provided to bcr/abl for it to transform fibroblasts.
Assuntos
Transformação Celular Neoplásica , Leucemia Mieloide/genética , Proteínas de Neoplasias/genética , Proteínas Oncogênicas Virais/fisiologia , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes/genética , Proteínas Virais/fisiologia , Vírus da Leucemia Murina de Abelson/fisiologia , Animais , Linhagem Celular , Transformação Celular Viral , Epitopos , Fibroblastos/patologia , Proteínas de Fusão bcr-abl , Produtos do Gene gag , Proteínas de Neoplasias/fisiologia , Proteínas Recombinantes de Fusão/fisiologia , Proteínas dos Retroviridae/fisiologia , TransfecçãoRESUMO
Chronic myelogenous leukemia (CML) is a human disease associated with a consistent chromosomal translocation that results in sequences from the c-abl locus on chromosome 9 being fused to sequences in a breakpoint cluster region (bcr) on chromosome 22. CML cells have two novel products: an 8.5-kilobase RNA transcript containing both abl and bcr and a 210-kilodalton phosphoprotein (P210) recognized by v-abl-specific antisera. To test whether the P210 is the product of the novel 8.5-kilobase bcr/abl fusion transcript, antibodies were prepared against c-abl and bcr determinants. By using these reagents and v-abl-specific antisera, it was demonstrated that the P210 in CML cells is indeed the protein product of the 8.5-kilobase transcript. By analogy to the gag/abl fusion protein of Abelson murine leukemia virus, the replacement of amino terminal c-abl sequences by bcr sequences in P210 may create a transforming protein involved in CML. A 190-kilodalton phosphoprotein that is a candidate for the normal bcr protein was identified in both HeLa and K562 cells.
Assuntos
Leucemia Mieloide/genética , Proteínas de Neoplasias/genética , Oncogenes , Vírus da Leucemia Murina de Abelson/genética , Animais , Cromossomos Humanos 21-22 e Y , Cromossomos Humanos 6-12 e X , Células HeLa/metabolismo , Humanos , Soros Imunes/imunologia , Cromossomo Filadélfia , Proteínas Tirosina Quinases/genética , Coelhos/imunologiaRESUMO
A phosphoinositide kinase specific for the D-3 position of the inositol ring, phosphatidylinositol (PI) 3-kinase, associates with activated receptors for platelet-derived growth factor, insulin, and colony-stimulating factor 1, with products of the oncogenes src, fms, yes, crk, and with polyomavirus middle T antigen. Efficient fibroblast transformation by proteins of the abl and src oncogene families requires activation of their protein-tyrosine kinase activity and membrane association via an amino-terminal myristoylation. We have demonstrated that the PI 3-kinase directly associates with autophosphorylated, activated protein-tyrosine kinase variants of the abl protein. In vivo, this association leads to accumulation of the highly phosphorylated products of PI 3-kinase, PI-3,4-bisphosphate and PI-3,4,5-trisphosphate, only in myristoylated, transforming abl protein variants. Myristoylation thus appears to be required to recruit PI 3-kinase activity to the plasma membrane for in vivo activation and correlates with the mitogenicity of the abl protein variants.
Assuntos
Transformação Celular Neoplásica , Genes abl , Variação Genética , Fosfotransferases/metabolismo , Animais , Linhagem Celular , Membrana Celular/enzimologia , Ativação Enzimática , Camundongos , Proteínas Oncogênicas v-abl/isolamento & purificação , Fosfatidilinositol 3-Quinases , Fosfotransferases/genética , Fosfotransferases/isolamento & purificaçãoRESUMO
A library of chicken genomic DNA was screened for sequences that could hybridize to a cloned DNA fragment containing the transforming gene (v-fps) of Fujinami sarcoma virus. In addition to c-fps, two unique chicken cellular DNA sequences were isolated that hybridized weakly to v-fps. These sequences hybridized with many other viral oncogenes encoding tyrosine kinases. Sequence analysis of the region where homology was detected revealed a region that is highly conserved among the tyrosine kinases both at the nucleotide and amino acid levels. Although we were unable to detect expression of either chicken cellular DNA sequence in a variety of avian tissues, the data suggest the existence of additional members of the tyrosine kinase gene family. Screening genomic libraries for sequences that hybridize weakly to functional regions of other genes may prove useful for the isolation and characterization of additional members of other gene families.
Assuntos
Vírus do Sarcoma Aviário/genética , Genes Virais , Genes , Oncogenes , Proteínas Tirosina Quinases/genética , Animais , Sequência de Bases , Transformação Celular Neoplásica , Galinhas , Clonagem Molecular , DNA/metabolismo , Hibridização de Ácido Nucleico , Homologia de Sequência do Ácido NucleicoRESUMO
N-terminal myristoylation can promote the association of proteins with the plasma membrane, a property that is required for oncogenic variants of Src and Abl to transform fibroblastic cell types. The P210bcr/abl protein of chronic myelogenous leukemia cells is not myristoylated and does not stably transform NIH 3T3 fibroblasts; however, it will transform lymphoid and myeloid cell types in vitro and in vivo, suggesting that myristoylation is not required for Abl variants to transform hematopoietic cells. To test this hypothesis, we introduced point mutations that disrupt myristoylation into two activated Abl proteins, v-Abl and a deletion mutant of c-Abl (delta XB), and examined their ability to transform an interleukin-3-dependent lymphoblastoid cell line, Ba/F3. Neither of the nonmyristoylated Abl proteins transformed NIH 3T3 fibroblasts, but like P210bcr/abl, both were capable of transforming the Ba/F3 cells to factor independence and tumorigenicity. Nonmyristoylated Abl variants did not associate with the plasma membrane in the transformed Ba/F3 cells. These results demonstrate that Abl proteins can transform hematopoietic cells in the absence of membrane association and suggest that distinct functions of Abl are required for transformation of fibroblast and hematopoietic cell types.
Assuntos
Transformação Celular Neoplásica , Ácidos Mirísticos/metabolismo , Proteínas Oncogênicas v-abl/genética , Processamento de Proteína Pós-Traducional , Células 3T3 , Animais , Transporte Biológico , Linhagem Celular , Análise Mutacional de DNA , Regulação Neoplásica da Expressão Gênica , Imuno-Histoquímica , Tecido Linfoide , Camundongos , Ácido Mirístico , Ácidos Mirísticos/isolamento & purificação , Proteínas Oncogênicas v-abl/isolamento & purificaçãoRESUMO
MYC is a major oncogenic driver of multiple myeloma (MM) and yet almost no therapeutic agents exist that target MYC in MM. Here we report that the let-7 biogenesis inhibitor LIN28B correlates with MYC expression in MM and is associated with adverse outcome. We also demonstrate that the LIN28B/let-7 axis modulates the expression of MYC, itself a let-7 target. Further, perturbation of the axis regulates the proliferation of MM cells in vivo in a xenograft tumor model. RNA-sequencing and gene set enrichment analyses of CRISPR-engineered cells further suggest that the LIN28/let-7 axis regulates MYC and cell cycle pathways in MM. We provide proof of principle for therapeutic regulation of MYC through let-7 with an LNA-GapmeR (locked nucleic acid-GapmeR) containing a let-7b mimic in vivo, demonstrating that high levels of let-7 expression repress tumor growth by regulating MYC expression. These findings reveal a novel mechanism of therapeutic targeting of MYC through the LIN28B/let-7 axis in MM that may impact other MYC-dependent cancers as well.
Assuntos
Regulação Neoplásica da Expressão Gênica , MicroRNAs/genética , Mieloma Múltiplo/genética , Interferência de RNA , Proteínas de Ligação a RNA/genética , Animais , Estudos de Casos e Controles , Ciclo Celular/genética , Linhagem Celular Tumoral , Análise por Conglomerados , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Genes myc , Xenoenxertos , Humanos , Camundongos , Mieloma Múltiplo/tratamento farmacológico , Mieloma Múltiplo/mortalidade , Mieloma Múltiplo/patologia , Prognóstico , Proteínas de Ligação a RNA/metabolismoRESUMO
The c-myc gene is frequently over-expressed in human cancers and is involved in regulation of proliferation, differentiation and apoptosis. c-Myc is a transcription factor that acts primarily by regulating the expression of other genes. However, it has been very difficult to identify bona fide c-Myc target genes that explain its diverse biological activities. The recent generation of c-myc deficient Rat1A fibroblasts with a profound and stable growth defect provides a new system to search for genes that can substitute for c-myc in proliferation. In this study, we have attempted to identify genes that rescue the slow growth phenotype of c-myc null cells through introduction of a series of potent cell cycle regulatory genes and several retroviral cDNA expression libraries. None of the candidate genes tested, including SV40 T-antigen and adenovirus E1A, caused reversal of the c-myc null growth defect. Furthermore, extensive screens with high-complexity retroviral cDNA libraries from three different tissue sources revealed that only c-myc and N-myc rescued the c-myc null slow-growth phenotype. Our data support the notion that there are no functional equivalents of the myc family of proto-oncogenes and also suggest that there are no c-Myc-activated genes that alone can substitute for c-Myc in control of cell proliferation.
Assuntos
Proteínas de Transporte , Proteínas de Ciclo Celular , Proteínas de Ligação a DNA , Fibroblastos/citologia , Proteínas Proto-Oncogênicas c-myc/fisiologia , Fatores de Transcrição/fisiologia , Proteínas E1A de Adenovirus/genética , Animais , Antígenos Transformantes de Poliomavirus/genética , Divisão Celular/genética , Linhagem Celular , Ciclina D1/genética , Ciclina E/genética , Fatores de Transcrição E2F , Fator de Transcrição E2F2 , Fator de Transcrição E2F3 , Sequências Hélice-Alça-Hélice , Humanos , Células K562 , Zíper de Leucina , Camundongos , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Ratos , Proteína 1 de Ligação ao Retinoblastoma , Fator de Transcrição DP1 , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
The Akt, Ras and STAT5 signaling pathways have each been linked to transformation of hematopoietic cells by BCR/ABL. However the relative contributions of these signaling pathways to BCR/ABL mediated cytokine-independent survival, proliferation and resistance to DNA damage-induced apoptosis have not been systematically defined. Here we report that activation of either Akt, Ras or STAT5 confers cytokine-independent survival to IL-3 dependent BaF3 cells. Ras or STAT5, but not Akt, also drives cytokine-independent proliferation and imparts sustained resistance to DNA damage-induced apoptosis. We also show that dominant negative (DN) inhibition of STAT5, but not Ras or Akt, significantly reduces resistance to DNA damage-induced apoptosis in BCR/ABL transformed BaF3 cells. Whereas inhibition of STAT5 or Ras alone does not compromise cytokine-independent proliferation of BaF3-BCR/ABL cells, simultaneous blockade of both STAT5 and Ras reduces proliferation and maximally sensitizes BaF3-BCR/ABL cells to DNA damage induced by gamma-irradiation, suggesting a cooperative role for these two signaling pathways in BCR/ABL transformation. The anti-apoptotic properties of BCR/ABL can be partly explained by an increase in the expression of Pim-1 and Bcl-XL, as ectopic expression of these STAT5 target genes imparts both cytokine-independent survival and partial gamma-radiation resistance. These data illustrate both cooperative and redundant effects of STAT5 and Ras signaling in BCR/ABL transformed cells, with STAT5 playing a dominant role in resistance to DNA damage-induced apoptosis.
Assuntos
Proteínas de Ligação a DNA/fisiologia , Proteínas de Fusão bcr-abl/fisiologia , Células-Tronco Hematopoéticas/fisiologia , Proteínas do Leite , Proteínas Serina-Treonina Quinases/fisiologia , Transativadores/fisiologia , Proteínas ras/fisiologia , Animais , Apoptose/fisiologia , Divisão Celular , Sobrevivência Celular/efeitos dos fármacos , Dano ao DNA , Proteínas de Ligação a DNA/genética , Proteínas de Fusão bcr-abl/genética , Vetores Genéticos/genética , Células-Tronco Hematopoéticas/efeitos dos fármacos , Interleucina-3/farmacologia , Cromossomo Filadélfia , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-akt , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteínas Proto-Oncogênicas c-pim-1 , Retroviridae , Fator de Transcrição STAT5 , Transdução de Sinais , Transativadores/genética , Transdução Genética , Proteína bcl-X , Proteínas ras/genéticaRESUMO
During differentiation in vitro, Embryonic Stem (ES) cells generate both primitive erythroid and definitive myeloid lineages in a process that mimics hematopoiesis in the mammalian yolk sac. To investigate leukemic transformation of these embryonic hematopoietic progenitors, we infected differentiating cultures of ES cells with the Chronic Myeloid Leukemia-specific BCR/ABL oncoprotein. Following a period of liquid culture, we isolated two transformed subclones, EB57 and EB67, that retained characteristics of embryonic hematopoietic progenitors and induced a fatal leukemia in mice characterized by massive splenomegaly and granulocytosis. Histopathology of the spleen revealed an abundance of undifferentiated blast-like cells. Investigation of the clonal origins of the granulocytes in the peripheral blood demonstrated that the injected donor cells contributed modestly to the granulocyte population while the majority were host-derived. EB57 secretes IL-3 and unidentified cytokines that can stimulate autocrine and paracrine cell proliferation, presumably accounting for the reactive granulocytosis in diseased mice. These BCR/ABL transformed hematopoietic derivatives of ES cells recapitulate the relationship of BCR/ABL expression to IL-3 production that has been described for primitive hematopoietic progenitors from human CML patients, and illustrates the potential for autocrine and paracrine effects of BCR/ABL-infected cells in murine models.
Assuntos
Transformação Celular Viral/genética , Proteínas de Fusão bcr-abl/fisiologia , Genes abl , Células-Tronco Hematopoéticas/patologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Animais , Antígenos de Superfície/biossíntese , Diferenciação Celular/fisiologia , Linhagem Celular Transformada , Células Precursoras Eritroides/citologia , Proteínas de Fusão bcr-abl/biossíntese , Proteínas de Fusão bcr-abl/genética , Granulócitos/patologia , Células-Tronco Hematopoéticas/fisiologia , Interleucina-3/biossíntese , Interleucina-3/fisiologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Camundongos , Camundongos Endogâmicos BALB C , Retroviridae/genética , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
BACKGROUND: Recent advances in high-throughput genomics technology have expanded our ability to catalogue allelic variants in large sets of candidate genes related to premature coronary artery disease. METHODS AND RESULTS: A total of 398 families were identified in 15 participating medical centers; they fulfilled the criteria of myocardial infarction, revascularization, or a significant coronary artery lesion diagnosed before 45 years in men or 50 years in women. A total of 62 vascular biology genes and 72 single-nucleotide polymorphisms were assessed. Previously undescribed variants in 3 related members of the thrombospondin protein family were prominent among a small set of single-nucleotide polymorphisms that showed a statistical association with premature coronary artery disease. A missense variant of thrombospondin 4 (A387P) showed the strongest association, with an adjusted odds ratio for myocardial infarction of 1.89 (P=0.002 adjusted for covariates) for individuals carrying the P allele. A variant in the 3' untranslated region of thrombospondin-2 (change of thymidine to guanine) seemed to have a protective effect against myocardial in individuals homozygous for the variant (adjusted odds ratio of 0.31; P=0.0018). A missense variant in thrombospondin-1 (N700S) was associated with an adjusted odds ratio for coronary artery disease of 11.90 (P=0.041) in homozygous individuals, who also had the lowest level of thrombospondin-1 by plasma assay (P=0.0019). CONCLUSIONS: This large-scale genetic study has identified the potential of multiple novel variants in the thrombospondin gene family to be associated with familial premature myocardial infarction. Notwithstanding multiple caveats, thrombospondins specifically and high-throughput genomic technology in general deserve further study in familial ischemic heart disease.
Assuntos
Doença da Artéria Coronariana/genética , Infarto do Miocárdio/diagnóstico , Infarto do Miocárdio/genética , Polimorfismo de Nucleotídeo Único/genética , Trombospondinas/genética , Adulto , Idade de Início , Alelos , Estudos de Casos e Controles , Angiografia Coronária , Doença da Artéria Coronariana/epidemiologia , Estenose Coronária/diagnóstico , Estenose Coronária/genética , Demografia , Feminino , Predisposição Genética para Doença , Testes Genéticos , Genótipo , Homozigoto , Humanos , Masculino , Metilenotetra-Hidrofolato Redutase (NADPH2) , Pessoa de Meia-Idade , Infarto do Miocárdio/epidemiologia , Razão de Chances , Oxirredutases atuantes sobre Doadores de Grupo CH-NH/genética , Valor Preditivo dos Testes , Trombospondina 1/genética , Estados UnidosRESUMO
The expression of the BCR-ABL fusion oncoprotein in primitive hematopoietic cells results in chronic myeloid leukemia. Over the past decade studies of several in vitro and in vivo cell systems revealed multiple signal transduction pathways activated by BCR-ABL. However, the precise function of BCR-ABL in the pathogenesis of CML is still unclear. The goal of this review is to synthesize data on intracellular signaling in the context of the diverse murine assay systems employed. We emphasize the importance of in vivo assays and assays using primary cells in understanding the biology of CML and the molecular mechanisms by which BCR-ABL exerts its effects.
Assuntos
Transformação Celular Neoplásica/genética , Proteínas de Fusão bcr-abl/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Animais , Bioensaio , Modelos Animais de Doenças , Células-Tronco Hematopoéticas/patologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Camundongos , Transdução de Sinais/genéticaRESUMO
Novel approaches have been designed to treat leukemia based on our understanding of the genetic and biochemical lesions present in different malignancies. This meeting report summarizes some of the recent advances in leukemia treatment. Based on the discoveries of cellular oncogenes, chromosomal translocations, monoclonal antibodies, multidrug resistance pumps, signal transduction pathways, genomics/proteonomic approaches to clinical diagnosis and mutations in biochemical pathways, clinicians and basic scientists have been able to identify the particular genetic mutations and signal transduction pathways involved as well as design more appropriate treatments for the leukemia patient. This meeting report discusses these exciting new therapies and the results obtained from ongoing clinical trials. Furthermore, rational approaches to treat complications of tumor lysis syndrome by administration of the recombinant urate oxidase protein, also known as rasburicase, which corrects the biochemical defect present in humans, were discussed. Clearly, over the past 25 years, molecular biology and biotechnology has provided the hematologist/oncologist novel bullets in their arsenal that will allow treatment by design in leukemia.
Assuntos
Antineoplásicos/uso terapêutico , Leucemia/tratamento farmacológico , Leucemia/fisiopatologia , Oncologia/tendências , HumanosRESUMO
Several environmental risk factors of cardiovascular disease are well established, but genetic risk alleles contributing to the disease in the general population are hotly debated. New strategies focusing on polymorphism discovery in candidate disease genes followed by tests of association to genes across the genome offer a pioneering approach to identifying risk alleles. Several hundred candidate genes for cardiovascular disease have been screened for common polymorphisms and these variants may provide susceptibility alleles which largely contribute to risk of cardiovascular disease in the general population. However, the impact of common susceptibility alleles for disease management will depend on many years of future investigation.
Assuntos
Coração/fisiologia , Miocárdio/metabolismo , Polimorfismo de Nucleotídeo Único/genética , Doenças Cardiovasculares/genética , Saúde da Família , Humanos , Polimorfismo Genético/genéticaRESUMO
A chronic myelogenous leukemia (CML)-like disease can be induced in mice by infecting hematopoietic stem cells with a BCR/ABL-containing retrovirus; serial transplantation produces either normal or leukemic animals. In many patients with CML, autografting produces transient Philadelphia chromosome (Ph)-negativity, but Ph-negative hematopoiesis is prolonged in some cases. These and other observations suggest that at diagnosis, CML patients may have substantial numbers of normal stem cells in their marrow, which may in certain circumstances regain a proliferative advantage if leukemic hematopoiesis can be suppressed by intensive chemotherapy. Thus autografting may have the capacity to restore normal hematopoiesis for long periods in patients not eligible for treatment by allogeneic bone marrow transplantation.
Assuntos
Transplante de Medula Óssea , Leucemia Mielogênica Crônica BCR-ABL Positiva/cirurgia , Animais , Modelos Animais de Doenças , Hematopoese/fisiologia , Células-Tronco Hematopoéticas/fisiologia , Humanos , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/fisiopatologia , Camundongos , Transplante AutólogoRESUMO
Mast cells (MCs) are critical components of the innate immune system and important for host defense, allergy, autoimmunity, tissue regeneration and tumor progression. Dysregulated MC development leads to systemic mastocytosis (SM), a clinically variable but often devastating family of hematologic disorders. Here we report that induced expression of Lin28, a heterochronic gene and pluripotency factor implicated in driving a fetal hematopoietic program, caused MC accumulation in adult mice in target organs such as the skin and peritoneal cavity. In vitro assays revealed a skewing of myeloid commitment in LIN28B-expressing hematopoietic progenitors, with increased levels of LIN28B in common myeloid and basophil-MC progenitors altering gene expression patterns to favor cell fate choices that enhanced MC specification. In addition, LIN28B-induced MCs appeared phenotypically and functionally immature, and in vitro assays suggested a slowing of MC terminal differentiation in the context of LIN28B upregulation. Finally, interrogation of human MC leukemia samples revealed upregulation of LIN28B in abnormal MCs from patients with SM. This work identifies Lin28 as a novel regulator of innate immune function and a new protein of interest in MC disease.