RESUMO
Simple retroviruses induce tumors by integrating into the host genome, activating cellular oncogenes and microRNAs, or inactivating tumor suppressor genes. The identification of these genes elucidates molecular mechanisms of tumorigenesis. In this study, we identified avian leukosis virus (ALV) proviral integration sites in rapid-onset B cell lymphomas arising <12 weeks after infection of chicken embryos. By using inverse PCR, 28 unique viral integration sites were identified in rapid-onset tumors. Integrations in the telomerase reverse transcriptase (TERT) promoter/enhancer region were observed in four different tumors, suggesting that this is a common integration site. These provirus integrations ranged from 217 to 2,584 bp upstream of the TERT transcription initiation site and were all in the opposite transcriptional orientation to TERT. Southern blots of tumor samples demonstrated that these integrations are clonal and therefore occurred early in the process of tumorigenesis. Real-time RT-PCR showed overexpression of TERT mRNA in tumors harboring viral integrations in the TERT promoter. Telomerase activity was also up-regulated in these tumors; however, telomere-length alterations were not detected. Furthermore, viral LTR sequences directly enhanced the expression of luciferase reporters containing the TERT promoter sequences. This study documents retroviral up-regulation of cellular TERT by insertional activation to initiate or enhance tumor progression.
Assuntos
Vírus da Leucose Aviária/genética , Leucose Aviária/virologia , Transformação Celular Viral/genética , Galinhas/virologia , Linfoma de Células B/veterinária , Mutagênese Insercional , Proteínas de Neoplasias/genética , Telomerase/genética , Integração Viral/genética , Animais , Leucose Aviária/genética , Embrião de Galinha , Galinhas/genética , Células Clonais/patologia , Células Clonais/virologia , Elementos Facilitadores Genéticos , Regulação Neoplásica da Expressão Gênica , Regulação Viral da Expressão Gênica , Genes myb , Linfoma de Células B/enzimologia , Linfoma de Células B/genética , Linfoma de Células B/virologia , Proteínas de Neoplasias/biossíntese , Regiões Promotoras Genéticas , Provírus/genética , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , RNA Neoplásico/biossíntese , RNA Neoplásico/genética , Telomerase/biossíntese , Sequências Repetidas Terminais , Regulação para CimaRESUMO
The avian leukosis virus DeltaLR-9 causes a high frequency of B-cell lymphomas within weeks after injection into 10-day-old chicken embryos. These lymphomas result from proviral integrations into the oncogene c-myb. In contrast, LR-9, which lacks the 42-nucleotide gag gene deletion of DeltaLR-9, does not cause a high frequency of c-myb-associated short-latency lymphomas. Although viral replication rates and spliced env mRNA levels were found to be similar for both viruses, DeltaLR-9 exhibited an increase in readthrough transcription compared to LR-9. The DeltaLR-9 deletion is located in the region of the gag gene corresponding to the matrix (MA) protein as well as in the negative regulator of splicing (NRS) element. To test whether disruption of the NRS or of the MA protein was responsible for inducing short-latency lymphomas, we generated viruses with NRS point mutations that maintained the wild-type Gag amino acid sequence. One of the mutant viruses induced an even higher incidence than DeltaLR-9 of short-latency lymphomas with viral integrations into c-myb. Thus, we propose that disruption of the NRS sequence promotes readthrough transcription and splicing to the downstream myb gene, causing overexpression of a slightly truncated Myb protein, which induces short-latency tumors.
Assuntos
Vírus da Leucose Aviária/genética , Vírus da Leucose Aviária/patogenicidade , Leucose Aviária/etiologia , Genes myb , Linfoma de Células B/etiologia , Animais , Leucose Aviária/genética , Leucose Aviária/virologia , Vírus da Leucose Aviária/fisiologia , Sequência de Bases , Embrião de Galinha , DNA Viral/genética , Genes env , Linfoma de Células B/genética , Linfoma de Células B/virologia , Proteínas Oncogênicas v-myb/genética , Proteínas Oncogênicas v-myb/fisiologia , Mutação Puntual , Processamento Pós-Transcricional do RNA/genética , Splicing de RNA/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Viral/genética , RNA Viral/metabolismo , Deleção de Sequência , Proteínas da Matriz Viral/genética , Proteínas da Matriz Viral/fisiologia , Integração Viral/genética , Replicação Viral/genéticaRESUMO
Avian retroviral integration into the c-myb locus is casually associated with the development of lymphomas in the bursa of Farbricius of chickens; these arise with a shorter latency than bursal lymphomas caused by deregulation of c-myc. This study indicates that c-myb mutation in embryonic bursal precursors leads to an oligoclonal population of developing bursal follicles, showing a variable propensity to form a novel lesion, the neoplastic follicle (NF). About half of such bursas rapidly developed lymphomas. Detection of changes in gene expression, during the development of neoplasms, was carried out by cDNA microarray analysis. The transcriptional signature of lymphomas with mutant c-myb was more limited than, and only partially shared with, those of bursal lymphomas caused by Myc or Rel oncogenes. The c-myb-associated lymphomas frequently showed overexpression of c-myc and altered expression of other genes involved in cell cycle control and proliferation-related signal transduction. Oligoclonal, NF-containing bursas lacked detectable c-myc overexpression and demonstrated a pattern of gene expression distinct from that of normal bursa and partially shared with the short-latency lymphomas. This functional genomic analysis uncovered several different pathways of lymphomagenesis by oncogenic transcription factors acting in a B-cell lineage.
Assuntos
Bolsa de Fabricius/patologia , Galinhas/genética , Regulação Neoplásica da Expressão Gênica/genética , Genes myb , Linfoma/genética , Animais , Vírus da Leucose Aviária/genética , Linfócitos B/metabolismo , Linfócitos B/patologia , Bolsa de Fabricius/metabolismo , Proteínas de Ciclo Celular/biossíntese , Proteínas de Ciclo Celular/genética , Divisão Celular/genética , Embrião de Galinha , Vírus Defeituosos/genética , Perfilação da Expressão Gênica , Genes myc , Genes rel , Linfoma/metabolismo , Linfoma/patologia , Metástase Neoplásica , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/genética , Análise de Sequência com Séries de Oligonucleotídeos , Fenótipo , Proteínas Proto-Oncogênicas c-myc/biossíntese , Proteínas Proto-Oncogênicas c-myc/genética , Transdução de Sinais/genética , Fatores de TempoRESUMO
Retroviruses require a balance of spliced and unspliced RNA for efficient replication. Here, we examined the effect of mutations in a splicing suppressor sequence called the negative regulator of splicing (NRS), located within the gag gene of Rous sarcoma virus. While the NRS mutant viruses showed only small changes in the levels of spliced env mRNAs, they had significant increases in src mRNA levels and transformed cells more efficiently than wild-type virus. None of these mutations prevented viral replication; however, some of the mutant viruses replicated more slowly than wild-type virus. In addition, increased transcriptional readthrough of the poly(A) site in the 3' LTR was observed with the NRS mutant viruses, suggesting that the wild-type NRS sequence promotes polyadenylation.
Assuntos
Vírus do Sarcoma Aviário/genética , Elementos Facilitadores Genéticos , Regulação Viral da Expressão Gênica , Genes src/genética , Poliadenilação , Splicing de RNA , Animais , Células Cultivadas , Embrião de Galinha , Fibroblastos , Genes gag , Mutação Puntual , RNA Mensageiro/metabolismo , RNA Viral/metabolismoRESUMO
The process of reproductive caste determination in eusocial insect colonies is generally understood to be mediated by environmental, rather than genetic factors. We present data demonstrating unexpected genetic differences between reproductive castes in a variant of the rough harvester ant, Pogonomyrmex rugosus var. fuscatus. Across multiple loci, queens were consistently more homozygous than expected, while workers were more heterozygous. Adult colony queens were divided into two highly divergent genetic groups, indicating the presence of two cryptic species, rather than a single population. The observed genetic differences between castes reflect differential representation of heterospecific and conspecific patrilines in these offspring groups. All workers were hybrids; by contrast, winged queens were nearly all pure-species. The complete lack of pure-species workers indicates a loss of worker potential in pure-species female offspring. Hybrids appear to be bipotential, but do not normally develop into reproductives because they are displaced by pure-species females in the reproductive pool. Genetic differences between reproductive castes are expected to be rare in non-hybridizing populations, but within hybrid zones they may be evolutionarily stable and thus much more likely to occur.