RESUMO
Haplotype-dependent allele-specific methylation (hap-ASM) can impact disease susceptibility, but maps of this phenomenon using stringent criteria in disease-relevant tissues remain sparse. Here we apply array-based and Methyl-Seq approaches to multiple human tissues and cell types, including brain, purified neurons and glia, T lymphocytes, and placenta, and identify 795 hap-ASM differentially methylated regions (DMRs) and 3,082 strong methylation quantitative trait loci (mQTLs), most not previously reported. More than half of these DMRs have cell type-restricted ASM, and among them are 188 hap-ASM DMRs and 933 mQTLs located near GWAS signals for immune and neurological disorders. Targeted bis-seq confirmed hap-ASM in 12/13 loci tested, including CCDC155, CD69, FRMD1, IRF1, KBTBD11, and S100A(∗)-ILF2, associated with immune phenotypes, MYT1L, PTPRN2, CMTM8 and CELF2, associated with neurological disorders, NGFR and HLA-DRB6, associated with both immunological and brain disorders, and ZFP57, a trans-acting regulator of genomic imprinting. Polymorphic CTCF and transcription factor (TF) binding sites were over-represented among hap-ASM DMRs and mQTLs, and analysis of the human data, supplemented by cross-species comparisons to macaques, indicated that CTCF and TF binding likelihood predicts the strength and direction of the allelic methylation asymmetry. These results show that hap-ASM is highly tissue specific; an important trans-acting regulator of genomic imprinting is regulated by this phenomenon; and variation in CTCF and TF binding sites is an underlying mechanism, and maps of hap-ASM and mQTLs reveal regulatory sequences underlying supra- and sub-threshold GWAS peaks in immunological and neurological disorders.
Assuntos
Metilação de DNA , Impressão Genômica , Haplótipos/genética , Polimorfismo de Nucleotídeo Único/genética , Locos de Características Quantitativas , Transativadores/genética , Alelos , Animais , Encéfalo/metabolismo , Encéfalo/patologia , Feminino , Estudo de Associação Genômica Ampla , Humanos , Doenças do Sistema Imunitário/genética , Macaca mulatta , Macaca radiata , Doenças do Sistema Nervoso/genética , Placenta/metabolismo , Placenta/patologia , Gravidez , Especificidade da Espécie , Linfócitos T/metabolismo , Linfócitos T/patologiaRESUMO
Familial cancer syndromes have revealed important fundamental features regarding how all cancers arise through destabilization of the genome, such that somatic evolution can select for the disruption of critical cellular coordinating and regulatory features. The authors examine those cellular genes and systems whose normal role is to preserve genomic integrity and relate them to the genetic foundations of heritable cancers. By examining how these cellular systems normally function, how family cancer genes are able to affect the process of tumor progression can be learned. In so doing, a clearer picture of how sporadic cancers arise is additionally gained.
Assuntos
Instabilidade Genômica/genética , Síndromes Neoplásicas Hereditárias/genética , Ciclo Celular , Dano ao DNA , Humanos , Fatores de RiscoRESUMO
TGFbeta is involved in the response to DNA damage and signaling the cell cycle checkpoint response, in large part achieved by modulating the activity of the ATM kinase. We have investigated if the presence of a common polymorphism in the TGFbeta receptor TGFBR1 might impact genomic instability in human colorectal cancer. In order to obtain statistically significant numbers of patients with the lesser polymorphism, 177 colorectal cancer patients were genotyped for either the major form of the TGFBR1 receptor gene, homozygous for an internal segment of 9 alanines (9A/9A), or the lesser form, heterozygous for the polymorphism containing 6 alanines (9A/6A). Intrachromosomal genomic instability in the tumors was then quantified by the robust inter-(simple sequence repeat) PCR method. Tumors from all 26 patients heterozygous with the (9A/6A) polymorphism in TGFBR1 exhibited significantly lower genomic instability than from a randomly selected set [the first identified] of 37 patients with the (9A/9A) polymorphism (p=0.0002, Mann-Whitney). The median age of onset for the (9A/6A) patients was 70 years, compared with a median age of onset of 63 years for the patients carrying the (9A/9A) form (p=0.031, Mann-Whitney). These results are consistent with the model wherein genomic instability facilitates tumor progression, with lesser instability associated with later disease presentation. Clinically, our findings may be developed into improved screening guidelines with respect to the age at which colonoscopy is initiated in carriers of the TGFBR1*6A allele.
Assuntos
Neoplasias Colorretais/genética , Repetições Minissatélites , Polimorfismo Genético , Proteínas Serina-Treonina Quinases/genética , Receptores de Fatores de Crescimento Transformadores beta/genética , Adulto , Idade de Início , Idoso , Idoso de 80 Anos ou mais , Alelos , Sequência de Bases , Dano ao DNA , Primers do DNA/genética , DNA de Neoplasias/genética , Feminino , Instabilidade Genômica , Heterozigoto , Homozigoto , Humanos , Masculino , Pessoa de Meia-Idade , Receptor do Fator de Crescimento Transformador beta Tipo I , Transdução de Sinais/genéticaRESUMO
In order to identify small regions of the genome whose specific copy number alteration is associated with high genomic instability in the form of overall genome-wide copy number aberrations, we have analyzed array-based comparative genomic hybridization (aCGH) data from 33 sporadic colorectal carcinomas. Copy number changes of a small number of specific regions were significantly correlated with elevated overall amplifications and deletions scattered throughout the entire genome. One significant region at 9q34 includes the c-ABL gene. Another region spanning 22q11-q13 includes the breakpoint cluster region (BCR) of the Philadelphia chromosome. Coordinate 22q11-q13 alterations were observed in 9 of 11 tumors with the 9q34 alteration. Additional regions on 1q and 14q were associated with overall genome-wide copy number changes, while copy number aberrations on chromosome 7p, 7q, and 13q21.1-q31.3 were found associated with this instability only in tumors from patients with a smoking history. Our analysis demonstrates there are a small number of regions of the genome where gain or loss is commonly associated with a tumor's overall level of copy number aberrations. Our finding BCR and ABL located within two of the instability-associated regions, and the involvement of these two regions occurring coordinately, suggests a system akin to the BCR-ABL translocation of CML may be involved in genomic instability in about one-third of human colorectal carcinomas.
Assuntos
Neoplasias Colorretais/genética , Dosagem de Genes , Genes abl , Instabilidade Genômica , Proteínas Proto-Oncogênicas c-bcr/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Cromossomos Artificiais Bacterianos/genética , Cromossomos Humanos Par 14/genética , Cromossomos Humanos Par 22/genética , Cromossomos Humanos Par 7/genética , Cromossomos Humanos Par 9/genética , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Hibridização de Ácido Nucleico , Análise de Sequência com Séries de OligonucleotídeosRESUMO
We have investigated transforming growth factor beta (TGF-beta)-mediated induction of actin stress fibers in normal and metastatic epithelial cells. We found that stress fiber formation requires de novo protein synthesis, p38Mapk and Smad signaling. We show that TGF-beta via Smad and p38Mapk up-regulates expression of actin-binding proteins including high-molecular-weight tropomyosins, alpha-actinin and calponin h2. We demonstrate that, among these proteins, tropomyosins are both necessary and sufficient for TGF-beta induction of stress fibers. Silencing of tropomyosins with short interfering RNAs (siRNAs) blocks stress fiber assembly, whereas ectopic expression of tropomyosins results in stress fibers. Ectopic-expression and siRNA experiments show that Smads mediate induction of tropomyosins and stress fibers. Interestingly, TGF-beta induction of stress fibers was not accompanied by changes in the levels of cofilin phosphorylation. TGF-beta induction of tropomyosins and stress fibers are significantly inhibited by Ras-ERK signaling in metastatic breast cancer cells. Inhibition of the Ras-ERK pathway restores TGF-beta induction of tropomyosins and stress fibers and thereby reduces cell motility. These results suggest that induction of tropomyosins and stress fibers play an essential role in TGF-beta control of cell motility, and the loss of this TGF-beta response is a critical step in the acquisition of metastatic phenotype by tumor cells.
Assuntos
Actinas/metabolismo , Movimento Celular/efeitos dos fármacos , Células Epiteliais/fisiologia , Fibras de Estresse/efeitos dos fármacos , Fator de Crescimento Transformador beta/farmacologia , Tropomiosina/metabolismo , Actinina/genética , Actinina/metabolismo , Animais , Proteínas de Ligação ao Cálcio , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Proteínas de Ligação a DNA/metabolismo , Células Epiteliais/citologia , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Regulação da Expressão Gênica , Humanos , Sistema de Sinalização das MAP Quinases/fisiologia , Camundongos , Proteínas dos Microfilamentos/genética , Proteínas dos Microfilamentos/metabolismo , Dados de Sequência Molecular , Metástase Neoplásica , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Proteínas Smad , Fibras de Estresse/metabolismo , Transativadores/metabolismo , Tropomiosina/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Proteínas ras/metabolismo , CalponinasRESUMO
BACKGROUND: Trisomy 21 causes Down syndrome (DS), but the mechanisms by which the extra chromosome leads to deficient intellectual and immune function are not well understood. RESULTS: Here, we profile CpG methylation in DS and control cerebral and cerebellar cortex of adults and cerebrum of fetuses. We purify neuronal and non-neuronal nuclei and T lymphocytes and find biologically relevant genes with DS-specific methylation (DS-DM) in each of these cell types. Some genes show brain-specific DS-DM, while others show stronger DS-DM in T cells. Both 5-methyl-cytosine and 5-hydroxy-methyl-cytosine contribute to the DS-DM. Thirty percent of genes with DS-DM in adult brain cells also show DS-DM in fetal brains, indicating early onset of these epigenetic changes, and we find early maturation of methylation patterns in DS brain and lymphocytes. Some, but not all, of the DS-DM genes show differential expression. DS-DM preferentially affected CpGs in or near specific transcription factor binding sites (TFBSs), implicating a mechanism involving altered TFBS occupancy. Methyl-seq of brain DNA from mouse models with sub-chromosomal duplications mimicking DS reveals partial but significant overlaps with human DS-DM and shows that multiple chromosome 21 genes contribute to the downstream epigenetic effects. CONCLUSIONS: These data point to novel biological mechanisms in DS and have general implications for trans effects of chromosomal duplications and aneuploidies on epigenetic patterning.
Assuntos
Aneuploidia , Encéfalo/metabolismo , Metilação de DNA/genética , Síndrome de Down/genética , Epigênese Genética , Adulto , Animais , Encéfalo/crescimento & desenvolvimento , Encéfalo/patologia , Cromossomos Humanos Par 21/genética , Ilhas de CpG/genética , Modelos Animais de Doenças , Síndrome de Down/patologia , Feto , Humanos , Camundongos , Linfócitos T/metabolismo , Linfócitos T/patologiaRESUMO
Human papillomavirus 16 (HPV-16) infection and tobacco use are associated with human oropharyngeal cancers. We conducted a study of the role of HPV and tobacco use in base of the tongue (BOT) cancers. DNA from 34 such cancers was subjected to HPV-16 and HPV-18-specific polymerase chain reaction analysis. Demographic and clinicopathologic data were obtained from each patient's medical record. HPV-16 was detected in 68% of tumors. Tobacco use was the only factor found to be significantly associated with HPV status. Tumors from 100% of patients who had never used tobacco tested positive for HPV, compared with only 56% of those who had ever used tobacco (Fisher exact test, p = 0.024). All tumors were associated with either tobacco use or HPV infection. These findings are consistent with the hypothesis that either tobacco use or HPV infection is necessary to the etiology of BOT tumors, and they suggest that tongue base carcinoma may be prevented by combining HPV vaccination with tobacco avoidance.
Assuntos
Carcinoma de Células Escamosas/etiologia , DNA Viral/análise , Papillomavirus Humano 16/isolamento & purificação , Papillomavirus Humano 18/isolamento & purificação , Infecções por Papillomavirus/epidemiologia , Fumar/efeitos adversos , Neoplasias da Língua/etiologia , Idoso , Idoso de 80 Anos ou mais , Carcinoma de Células Escamosas/virologia , Feminino , Papillomavirus Humano 16/genética , Papillomavirus Humano 18/genética , Humanos , Incidência , Masculino , Pessoa de Meia-Idade , Infecções por Papillomavirus/complicações , Fumar/epidemiologia , Neoplasias da Língua/virologiaRESUMO
Our previous allelotyping studies of 59 sporadic colorectal cancers revealed that loss of heterozygosity is most frequent for regions of chromosomes 14 and 18. Yet subsequent BAC microarray comparative genomic hybridization studies of the same tumor DNAs showed no corresponding pattern of copy number alteration for chromosome 14. To clarify this apparent discrepancy, we utilized hybridization to SNP microarrays; this revealed frequent uniparentalism for chromosome 14 and for chromosome 18. Based on the BAC array results combined with fluorescent in situ hybridization data, it was evident that uniparental disomy was occurring in many colorectal cancers as well as in additional chromosomes, and often coordinately involved chromosomes 14 and 18. Further studies examined the possibility that uniparentalism was directed towards the selection for imprinted genes, but no association with imprinting was observed.