RESUMO
The impact of genome organization on the control of gene expression persists as a major challenge in regulatory biology. Most efforts have focused on the role of CTCF-enriched boundary elements and TADs, which enable long-range DNA-DNA associations via loop extrusion processes. However, there is increasing evidence for long-range chromatin loops between promoters and distal enhancers formed through specific DNA sequences, including tethering elements, which bind the GAGA-associated factor (GAF). Previous studies showed that GAF possesses amyloid properties in vitro, bridging separate DNA molecules. In this study, we investigated whether GAF functions as a looping factor in Drosophila development. We employed Micro-C assays to examine the impact of defined GAF mutants on genome topology. These studies suggest that the N-terminal POZ/BTB oligomerization domain is important for long-range associations of distant GAGA-rich tethering elements, particularly those responsible for promoter-promoter interactions that coordinate the activities of distant paralogous genes.
Assuntos
Proteínas de Drosophila , Drosophila , Animais , Cromatina/genética , DNA/metabolismo , Proteínas de Ligação a DNA/genética , Drosophila/genética , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Elementos Facilitadores Genéticos , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Recently, covalent modifications of RNA, such as methylation, have emerged as key regulators of all aspects of RNA biology and have been implicated in numerous diseases, for instance, cancer. Here, we undertook a combination of in vitro and in vivo screens to test 78 potential methyltransferases for their roles in hepatocellular carcinoma (HCC) cell proliferation. We identified methyltransferase-like protein 6 (METTL6) as a crucial regulator of tumor cell growth. We show that METTL6 is a bona fide transfer RNA (tRNA) methyltransferase, catalyzing the formation of 3-methylcytidine at C32 of specific serine tRNA isoacceptors. Deletion of Mettl6 in mouse stem cells results in changes in ribosome occupancy and RNA levels, as well as impaired pluripotency. In mice, Mettl6 knockout results in reduced energy expenditure. We reveal a previously unknown pathway in the maintenance of translation efficiency with a role in maintaining stem cell self-renewal, as well as impacting tumor cell growth profoundly.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Animais , Carcinoma Hepatocelular/genética , Proliferação de Células , Neoplasias Hepáticas/genética , Metiltransferases/genética , Metiltransferases/metabolismo , Camundongos , RNA , RNA de Transferência/genética , RNA de Transferência/metabolismo , tRNA MetiltransferasesRESUMO
After fertilization, to initiate development, gametes are reprogramed to become totipotent. Approximately half of the mammalian genome consists of repetitive elements, including retrotransposons, some of which are transcribed after fertilization. Retrotransposon activation is generally assumed to be a side effect of the extensive chromatin remodeling underlying the epigenetic reprogramming of gametes. Here, we used a targeted epigenomic approach to address whether specific retrotransposon families play a direct role in chromatin organization and developmental progression. We demonstrate that premature silencing of LINE-1 elements decreases chromatin accessibility, whereas prolonged activation prevents the gradual chromatin compaction that occurs naturally in developmental progression. Preventing LINE-1 activation and interfering with its silencing decreases developmental rates independently of the coding nature of the LINE-1 transcript, thus suggesting that LINE-1 functions primarily at the chromatin level. Our data suggest that activation of LINE-1 regulates global chromatin accessibility at the beginning of development and indicate that retrotransposon activation is integral to the developmental program.
Assuntos
Blástula/metabolismo , Montagem e Desmontagem da Cromatina/genética , Desenvolvimento Embrionário/genética , Regulação da Expressão Gênica no Desenvolvimento , Elementos Nucleotídeos Longos e Dispersos/fisiologia , Zigoto/metabolismo , Animais , Cruzamentos Genéticos , Técnicas de Cultura Embrionária , Feminino , Fertilização , Inativação Gênica , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Técnicas Analíticas Microfluídicas , RNA Mensageiro/genética , Nucleases dos Efetores Semelhantes a Ativadores de Transcrição , Transcrição GênicaRESUMO
Transvection, a chromosome pairing-dependent form of trans-based gene regulation, is potentially widespread in the Drosophila melanogaster genome and varies across cell types and within tissues in D. melanogaster, characteristics of a complex trait. Here, we demonstrate that the trans-interactions at the Malic enzyme (Men) locus are, in fact, transvection as classically defined and are plastic with respect to both genetic background and environment. Using chromosomal inversions, we show that trans-interactions at the Men locus are eliminated by changes in chromosomal architecture that presumably disrupt somatic pairing. We further show that the magnitude of transvection at the Men locus is modified by both genetic background and environment (temperature), demonstrating that transvection is a plastic phenotype. Our results suggest that transvection effects in D. melanogaster are shaped by a dynamic interplay between environment and genetic background. Interestingly, we find that cis-based regulation of the Men gene is more robust to genetic background and environment than trans-based. Finally, we begin to uncover the nonlocal factors that may contribute to variation in transvection overall, implicating Abd-B in the regulation of Men in cis and in trans in an allele-specific and tissue-specific manner, driven by differences in expression of the two genes across genetic backgrounds and environmental conditions.
