RESUMEN

There is growing evidence that peptides encoded by small open-reading frames (sORF or smORF) can fulfill various cellular functions and define a novel class regulatory molecules. To which extend transcripts encoding only smORF peptides compare with canonical protein-coding genes, yet remain poorly understood. In particular, little is known on whether and how smORF-encoding RNAs might need tightly regulated expression within a given tissue, at a given time during development. We addressed these questions through the analysis of Drosophila polished rice (pri, a.k.a. tarsal less or mille pattes), which encodes four smORF peptides (11-32 amino acids in length) required at several stages of development. Previous work has shown that the expression of pri during epidermal development is regulated in the response to ecdysone, the major steroid hormone in insects. Here, we show that pri transcription is strongly upregulated by ecdysone across a large panel of cell types, suggesting that pri is a core component of ecdysone response. Although pri is produced as an intron-less short transcript (1.5 kb), genetic assays reveal that the developmental functions of pri require an unexpectedly large array of enhancers (spanning over 50 kb), driving a variety of spatiotemporal patterns of pri expression across developing tissues. Furthermore, we found that separate pri enhancers are directly activated by the ecdysone nuclear receptor (EcR) and display distinct regulatory modes between developmental tissues and/or stages. Alike major developmental genes, the expression of pri in a given tissue often involves several enhancers driving apparently redundant (or shadow) expression, while individual pri enhancers can harbor pleiotropic functions across tissues. Taken together, these data reveal the broad role of Pri smORF peptides in ecdysone signaling and show that the cis-regulatory architecture of the pri gene contributes to shape distinct spatial and temporal patterns of ecdysone response throughout development.

RESUMEN

In many animals, progression of developmental stages is temporally controlled by steroid hormones. In Drosophila, the level of ecdysone titer oscillates and developmental stage transitions, such as larval molting and metamorphosis, are induced at each of ecdysone peaks. Ecdysone titer also peaks at the stage of mid-embryogenesis and the embryonic ecdysone is necessary for morphogenesis of several organs, although the regulatory mechanisms of embryonic organogenesis dependent on ecdysone signaling are still open questions. In this study, we find that absence or interruption of embryonic ecdysone signaling caused multiple defects in the tracheal system, including decrease in luminal protein deposition, uneven dilation of the dorsal trunk and loss of terminal branches. We also reveal that an ecdysone-inducible gene polished rice (pri) is essential for tip cell fate decision in dorsal branches. As over-expression of pri can restore the defects caused by disturbance of ecdysone biosynthesis, pri functions as one of the major mediators of embryonic ecdysone signal in tracheogenesis. These results demonstrate that ecdysone and its downstream target pri play essential roles in tracheal development by modulating cell fate decision.

Asunto(s)

Proteínas de Drosophila/metabolismo , Drosophila melanogaster/embriología , Drosophila melanogaster/metabolismo , Ecdisona/metabolismo , Embrión no Mamífero/metabolismo , Organogénesis , Transaldolasa/metabolismo , Animales , Diferenciación Celular , Linaje de la Célula , Proteínas de Drosophila/genética , Regulación del Desarrollo de la Expresión Génica , Modelos Biológicos , Mutación/genética , Fenotipo , Tráquea/citología , Tráquea/embriología , Tráquea/metabolismo , Transaldolasa/genética

RESUMEN

To compensate for accumulating damages and cell death, adult homeostasis (e.g., body fluids and secretion) requires organ regeneration, operated by long-lived stem cells. How stem cells can survive throughout the animal life remains poorly understood. Here we show that the transcription factor Shavenbaby (Svb, OvoL in vertebrates) is expressed in renal/nephric stem cells (RNSCs) of Drosophila and required for their maintenance during adulthood. As recently shown in embryos, Svb function in adult RNSCs further needs a post-translational processing mediated by the Polished rice (Pri) smORF peptides and impairing Svb function leads to RNSC apoptosis. We show that Svb interacts both genetically and physically with Yorkie (YAP/TAZ in vertebrates), a nuclear effector of the Hippo pathway, to activate the expression of the inhibitor of apoptosis DIAP1. These data therefore identify Svb as a nuclear effector in the Hippo pathway, critical for the survival of adult somatic stem cells.

Asunto(s)

Células Madre Adultas/metabolismo , Proteínas de Unión al ADN/metabolismo , Proteínas de Drosophila/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Transactivadores/metabolismo , Factores de Transcripción/metabolismo , Animales , Apoptosis/genética , Apoptosis/fisiología , Proteínas de Unión al ADN/genética , Drosophila , Proteínas de Drosophila/genética , Etiquetado Corte-Fin in Situ , Péptidos y Proteínas de Señalización Intracelular/genética , Proteínas Nucleares/genética , Proteínas Serina-Treonina Quinasas/genética , Transducción de Señal/genética , Transducción de Señal/fisiología , Transactivadores/genética , Factores de Transcripción/genética , Proteínas Señalizadoras YAP

RESUMEN

Animal development fundamentally relies on the precise control, in space and time, of genome expression. Whereas we have a wealth of information about spatial patterning, the mechanisms underlying temporal control remain poorly understood. Here we show that Pri peptides, encoded by small open reading frames, are direct mediators of the steroid hormone ecdysone for the timing of developmental programs in Drosophila. We identify a previously uncharacterized enzyme of ecdysone biosynthesis, GstE14, and find that ecdysone triggers pri expression to define the onset of epidermal trichome development, through post-translational control of the Shavenbaby transcription factor. We show that manipulating pri expression is sufficient to either put on hold or induce premature differentiation of trichomes. Furthermore, we find that ecdysone-dependent regulation of pri is not restricted to epidermis and occurs over various tissues and times. Together, these findings provide a molecular framework to explain how systemic hormonal control coordinates specific programs of differentiation with developmental timing.

Asunto(s)

Arrestinas/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Ecdisona/metabolismo , Regulación del Desarrollo de la Expresión Génica/fisiología , Glutatión Transferasa/metabolismo , Receptores de Esteroides/metabolismo , Animales , Arrestinas/genética , Diferenciación Celular/genética , Proteínas de Drosophila/genética , Ecdisona/genética , Glutatión Transferasa/genética , Mutación/genética , Receptores de Esteroides/genética , Transducción de Señal/fisiología , Transaldolasa/genética , Transaldolasa/metabolismo

RESUMEN

Systematic transcriptome analysis has revealed that a vast majority of the mammalian genome is transcribed into RNA, thus establishing the concept of "pervasive transcription." More than half of these RNAs do not encode proteins, and they are collectively called noncoding RNAs. Although the physiological relevance of the transcription of these noncoding RNAs has remained unclear, it was recently proposed that one of the major roles of long noncoding RNAs (lncRNAs) in the nucleus is the regulation of gene expression at the transcriptional level via histone or DNA modification. In this review, we will summarize the advancement of our understanding of the molecular mechanisms of lncRNAs. This article is part of a Special Issue entitled: Chromatin and epigenetic regulation of animal development.

Asunto(s)

Cromatina/metabolismo , Epigénesis Genética/fisiología , ARN Largo no Codificante/biosíntesis , Transcripción Genética/fisiología , Animales , Cromatina/genética , Histonas/genética , Histonas/metabolismo , Humanos , Procesamiento Proteico-Postraduccional/fisiología , ARN Largo no Codificante/genética

RESUMEN

Nuclear receptor transcription factor family members share target sequence similarity; however, little is known about how these factors exert their specific regulatory control. Here, we examine the mechanism regulating the expression of the Drosophila EDG84A gene, a target gene of the orphan nuclear receptor ßFTZ-F1, as a model to study the cooperative behavior among nuclear receptors. We show that the three nuclear receptors ßFTZ-F1, DHR3, and DHR39 bind to a common element in the EDG84A promoter. The expression level of the EDG84A promoter-lacZ reporter genes in DHR39-induced and mutant animals, respectively, suggests that DHR39 works as a repressor. The activity of a reporter gene carrying a mutation preventing DHR3 binding was reduced in ftz-f1 mutants and rescued by the induced expression of ßFTZ-F1, suggesting that DHR3 and ßFTZ-F1 activate the EDG84A gene in a redundant manner. A reporter gene carrying a mutation that abolishes DHR39 and FTZ-F1 binding was prematurely expressed, and the expression level of the reporter gene carrying a mutation preventing DHR3 binding was reduced. These findings suggest that the temporal expression of this gene is mainly controlled by ßFTZ-F1 but that the binding of DHR3 is also important. Comparison of the binding site sequence among Drosophila species suggests that DHR3 binding ability was gained after the melanogaster subgroup evolved, and this ability may contribute to the robust expression of this gene. These results show the complicated regulatory mechanisms utilized by multiple nuclear receptors to properly regulate the expression of their target gene through a single target site.

Asunto(s)

Proteínas de Unión al ADN/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Proteínas de Insectos/genética , Receptores Citoplasmáticos y Nucleares/metabolismo , Receptores de Esteroides/metabolismo , Animales , Sitios de Unión , Proteínas de Unión al ADN/genética , Proteínas de Drosophila/genética , Drosophila melanogaster/crecimiento & desarrollo , Drosophila melanogaster/metabolismo , Regulación del Desarrollo de la Expresión Génica/genética , Genes Reporteros , Proteínas de Insectos/metabolismo , Metamorfosis Biológica , Mutación , Regiones Promotoras Genéticas/genética , Receptores Citoplasmáticos y Nucleares/genética , Receptores de Esteroides/genética , Activación Transcripcional/genética

RESUMEN

Genome analysis has identified a number of putative non-protein-coding transcripts that do not contain ORFs longer than 100 codons. Although evidence strongly suggests that non-coding RNAs are important in a variety of biological phenomena, the discovery of small peptide-coding mRNAs confirms that some transcripts that have been assumed to be non-coding actually have coding potential. Their abundance and importance in biological phenomena makes the sorting of non-coding RNAs from small peptide-coding mRNAs a key issue in functional genomics. However, validating the coding potential of small peptide-coding RNAs is complicated, because their ORF sequences are usually too short for computational analysis. In this review, we discuss computational and experimental methods for validating the translatability of these non-coding RNAs.

Asunto(s)

Sistemas de Lectura Abierta/genética , Biosíntesis de Proteínas/genética , ARN no Traducido/genética , ARN no Traducido/metabolismo , Transcripción Genética , Empalme Alternativo/genética , Animales , Biología Computacional , Regulación de la Expresión Génica , Humanos , Redes y Vías Metabólicas/genética , Isoformas de ARN/genética

Asunto(s)

Drosophila/genética , ARN Mensajero , ARN no Traducido , Animales , Perfilación de la Expresión Génica , Peso Molecular , Péptidos/genética , Proteínas/genética , Transcripción Genética/genética

RESUMEN

Generally, bioactive small peptides are derived from precursors with signal sequences at their N-terminal ends, which undergo modification and proteolysis through a secretory pathway. By contrast, small peptides encoded in short open reading frames (sORF) lack signaling sequences and therefore are released into the cytoplasm, which may result in their having functions distinct from those of secreted peptides. Several small peptides encoded by sORF are involved in the morphogenesis of multicellular organisms. POLARIS, ROTUNDIFOLIA4, and Enod40 are plant peptides that are involved, respectively, in root formation, leaf shape control, and cortical cell division during nodule formation. Brick1/HSPC300 is an evolutionarily conserved component of the actin reorganization complex. polished rice/tarsal-less and mille-pattes encode related small peptides that are required for epithelial morphogenesis in Drosophila and segmentation in Tribolium. There are only a few known examples of small peptides encoded by sORF, and their molecular functions are still largely obscure. Nevertheless, an increasing number of sORF genes is being identified, and further research should reveal their roles in novel molecular mechanisms underlying developmental events.

Asunto(s)

Biología Evolutiva/métodos , Sistemas de Lectura Abierta , Péptidos/química , ARN no Traducido/química , Animales , Arabidopsis , Codón , Drosophila , Regulación del Desarrollo de la Expresión Génica , Genes , Genes de Plantas , Proteínas de Plantas , Estructura Terciaria de Proteína

RESUMEN

Transcription levels of the genes on X chromosome are regulated through the dosage compensation mechanisms. The dosage compensation complex (DCC) localizes to X chromosome and activates the transcription of target genes in male 2-fold more than in female. Drosophila maleless (MLE), an ATPase/helicase, is a component of the DCC and essential for the viability of male flies. However, the functions of MLE on gene expression are not clear. RNA helicase A (RHA) is a homologue of Drosophila MLE and mediates the expression of several genes. RHA recruits preinitiation complex via the minimal transactivation domain (MTAD), consisting of 50 amino acids to target promoters. The tryptophan residues in MTAD are important for transactivation via RHA. The amino acid sequence of MTAD is conserved in MLE. In this study, we assessed whether the functions of MTAD are conserved in fruit fly by investigating the transcriptional activity of MLE. Transactivation assay indicated the MTAD of MLE had transcriptional activity in Schneider's cells. In vitro binding assays revealed that MLE recruited RNA polymerase II (Pol II) complexes through MTAD. Reporter assays showed that the MTAD, especially tryptophan residues, is important for transcription from roX promoter, similar to RHA. The results confirm that MTAD of MLE mediates the expression of MLE target genes through recruitment of Pol II.

Asunto(s)

Proteínas Cromosómicas no Histona/genética , Proteínas Cromosómicas no Histona/metabolismo , ADN Helicasas/genética , ADN Helicasas/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila/genética , Drosophila/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Activación Transcripcional , Animales , Animales Modificados Genéticamente , Línea Celular , Proteínas Cromosómicas no Histona/química , ADN Helicasas/química , Compensación de Dosificación (Genética) , Proteínas de Drosophila/química , Femenino , Genes de Insecto , Masculino , Modelos Biológicos , Estructura Terciaria de Proteína , ARN Helicasas/química , ARN Helicasas/genética , ARN Helicasas/metabolismo , ARN Polimerasa II/metabolismo , Factores de Transcripción/química , Cromosoma X/genética

Asunto(s)

Sistemas de Lectura Abierta/genética , Péptidos/genética , Péptidos/fisiología , Animales , Proteínas de Drosophila , Desarrollo Embrionario/genética , Perfilación de la Expresión Génica , Péptidos/química , ARN no Traducido/genética , ARN no Traducido/fisiología , Transaldolasa

RESUMEN

Regulatory mechanisms controlling the timing of developmental events are crucial for proper development to occur. ftz-f1 is expressed in a temporally regulated manner following pulses of ecdysteroid and this precise expression is necessary for the development of Drosophila melanogaster. To understand how insect hormone ecdysteroids regulate the timing of FTZ-F1 expression, we purified a DNA binding regulator of ftz-f1. Mass spectroscopy analysis revealed this protein to be a fly homolog of mammalian B lymphocyte-induced maturation protein 1 (Blimp-1). Drosophila Blimp-1 (dBlimp-1) is induced directly by 20-hydroxyecdysone, and its product exists during high-ecdysteroid periods and turns over rapidly. Forced expression of dBlimp-1 and RNA interference analysis indicate that dBlimp-1 acts as a repressor and controls the timing of FTZ-F1 expression. Furthermore, its prolonged expression results in delay of pupation timing. These results suggest that the transient transcriptional repressor dBlimp-1 is important for determining developmental timing in the ecdysone-induced pathway.

Asunto(s)

Proteínas de Drosophila/metabolismo , Drosophila melanogaster/efectos de los fármacos , Drosophila melanogaster/embriología , Ecdisona/farmacología , Proteínas Represoras/metabolismo , Transcripción Genética , Animales , Sitios de Unión , Extractos Celulares , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/aislamiento & purificación , Embrión no Mamífero/efectos de los fármacos , Embrión no Mamífero/metabolismo , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Regiones Promotoras Genéticas/genética , Pupa/efectos de los fármacos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Proteínas Represoras/genética , Proteínas Represoras/aislamiento & purificación , Homología de Secuencia de Aminoácido , Termodinámica , Factores de Tiempo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Transcripción Genética/efectos de los fármacos

RESUMEN

Transcriptome analyses in eukaryotes, including mice and humans, have identified polyA-containing transcripts that lack long open reading frames (ORFs; >100 amino acids). These transcripts are believed most likely to function as non-coding RNAs, but their translational capacities and biological activities have not been characterized in detail. Here, we report that polished rice (pri), which was previously identified as a gene for a non-coding RNA in Drosophila, is in fact transcribed into a polycistronic mRNA that contains evolutionarily conserved short ORFs that encode 11 or 32 amino acid-long peptides. pri was expressed in all epithelial tissues during embryogenesis. The loss of pri function completely eliminated apical cuticular structures, including the epidermal denticles and tracheal taenidia, and also caused defective tracheal-tube expansion. We found that pri is essential for the formation of specific F-actin bundles that prefigures the formation of the denticles and taenidium. We provide evidences that pri acts non-cell autonomously and that four of the conserved pri ORFs are functionally redundant. These results demonstrate that pri has essential roles in epithelial morphogenesis by regulating F-actin organization.

Asunto(s)

Actinas/metabolismo , Drosophila/embriología , Desarrollo Embrionario/genética , Epitelio/embriología , Péptidos/metabolismo , ARN Mensajero/genética , Animales , Secuencia de Bases , Diferenciación Celular/genética , Secuencia Conservada/genética , Drosophila/citología , Drosophila/metabolismo , Epitelio/metabolismo , Evolución Molecular , Regulación del Desarrollo de la Expresión Génica/genética , Genes/genética , Datos de Secuencia Molecular , Sistemas de Lectura Abierta/genética , Péptidos/genética , Homología de Secuencia de Ácido Nucleico

RESUMEN

RNAi is a gene-silencing phenomenon mediated by double-stranded RNA (dsRNA) and has become a powerful tool to elucidate gene function. To accomplish rapid construction of transgenes expressing dsRNA in Drosophila, we developed a novel transformation vector, pRISE, which contains an inverted repeat of the attR1-ccdB-attR2 cassette for in vitro recombination and a pentameric GAL4 binding site for conditional expression. These features enabled us to construct RNAi transgenes without a complicated cloning scheme. In cultured cells and transgenic flies, pRISE constructs carrying dsRNA transgenes induced effective RNAi against an EGFP transgene and the endogenous white gene, respectively. These results indicate that pRISE is a convenient transformation vector for studies of multiple Drosophila genes for which functional information is lacking.

Asunto(s)

Elementos Transponibles de ADN/genética , Drosophila/genética , Vectores Genéticos/síntesis química , ARN Interferente Pequeño/genética , Transgenes , Transportadoras de Casetes de Unión a ATP/metabolismo , Animales , Animales Modificados Genéticamente , Células Cultivadas , Proteínas de Drosophila/deficiencia , Proteínas de Drosophila/metabolismo , Proteínas del Ojo/metabolismo , Interferencia de ARN , Recombinación Genética , Transformación Genética

RESUMEN

Maternally supplied germ plasm is essential for germ lineage establishment in many species, but the molecular details are still largely unknown, especially in vertebrates, and identification of novel factors that localize to germ plasm is desirable. We previously reported that one of the components of zebrafish germ plasm is mRNA of the bruno-like (brul) gene, a homologue of bruno, which, in Drosophila, is known to participate in germ lineage establishment. Here, we show that not only mRNA but also protein of brul is localized to the zebrafish germ plasm at the ends of the cleavage furrows. In 4- and 8-cell stage embryos, Brul protein is localized to the periphery of the blastomeres, as well as to the ends of the cleavage furrows, forming numerous minute particles. These particles appear at the cortex of the fertilized egg within 10 min after fertilization. Surprisingly, these distinctive localizations, as well as the minute particles, completely disappeared by the 16-cell stage, although relatively weak expression was detected ubiquitously throughout embryogenesis. This is the first report of a protein that localizes to the germ plasm in zebrafish.

Asunto(s)

Proteínas de Unión al ARN/metabolismo , Proteínas de Pez Cebra/metabolismo , Pez Cebra/embriología , Pez Cebra/metabolismo , Animales , Blastocisto/metabolismo , Proteínas CELF1 , Fase de Segmentación del Huevo/metabolismo , Femenino , Células Germinativas/metabolismo , Inmunohistoquímica , Masculino

Asunto(s)

MicroARNs/fisiología , ARN no Traducido/fisiología , Animales , Células Cultivadas , Dendritas/genética , Dendritas/fisiología , Regulación del Desarrollo de la Expresión Génica , Proteínas de Homeodominio/genética , Humanos , Modelos Genéticos , ARN Largo no Codificante

RESUMEN

One of the most surprising results to emerge from mammalian cDNA sequencing projects is that thousands of mRNA-like non-coding RNAs (ncRNAs) are expressed and constitute at least 10% of poly(A)(+) RNAs. In most cases, however, the functions of these RNA molecules remain unclear. To clarify the biological significance of mRNA-like ncRNAs, we computationally screened 11,691 Drosophila melanogaster full-length cDNAs. After eliminating presumable protein-coding transcripts, 136 were identified as strong candidates for mRNA-like ncRNAs. Although most of these putative ncRNAs are found throughout the Drosophila genus, predicted amino acid sequences are not conserved even in related species, suggesting that these transcripts are actually non-coding RNAs. In situ hybridization analyses revealed that 35 of the transcripts are expressed during embryogenesis, of which 27 were detected only in specific tissues including the tracheal system, midgut primordial cells, visceral mesoderm, germ cells and the central and peripheral nervous system. These highly regulated expression patterns suggest that many mRNA-like ncRNAs play important roles in multiple steps of organogenesis and cell differentiation in Drosophila. This is the first report that the majority of mRNA-like ncRNAs in a model organism are expressed in specific tissues and cell types.

Asunto(s)

Drosophila/genética , ARN Mensajero/genética , ARN no Traducido/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Diferenciación Celular/genética , Secuencia Conservada , ADN Complementario/análisis , ADN Complementario/genética , Drosophila/embriología , Desarrollo Embrionario/genética , Evolución Molecular , Regulación del Desarrollo de la Expresión Génica , Modelos Genéticos , Sistemas de Lectura Abierta/genética , Organogénesis/genética , ARN Mensajero/química , ARN no Traducido/química , Especificidad de la Especie , Transcripción Genética

Asunto(s)

Epigénesis Genética/genética , Regulación del Desarrollo de la Expresión Génica/genética , ARN no Traducido/fisiología , Transcripción Genética/genética , Animales , Ensamble y Desensamble de Cromatina/genética , Compensación de Dosificación (Genética) , Reordenamiento Génico/genética , Genoma , Heterocromatina/genética , Humanos , Interferencia de ARN/fisiología

RESUMEN

Members of the maf gene family encode basic/leucine zipper transcription factors and play important roles during cell differentiation and organogenesis in vertebrate development. In this study, we show that the maf family is evolutionarily conserved and that the Drosophila maf (Dmaf) gene is expressed in somatic gonadal cells. During embryonic development, Dmaf mRNA is detected in somatic gonadal precursor cells emerging from dorsolateral mesoderm. Relatively weak expression is also observed in subset of neuronal cells in the central nervous system. In adult flies, Dmaf is expressed in somatic gonadal cells surrounding developing oocytes and spermatocytes. These results suggest a specific function for Dmaf in gonadal development, including migration and differentiation of primordial germ cells.

Asunto(s)

Proteínas Bacterianas/metabolismo , Drosophila/embriología , Gónadas/metabolismo , Factores de Transcripción/metabolismo , Secuencia de Aminoácidos , Animales , Tipificación del Cuerpo , Drosophila/genética , Evolución Molecular , Células Germinativas , Gónadas/citología , Gónadas/embriología , Datos de Secuencia Molecular , Familia de Multigenes , Filogenia , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Factores de Transcripción/clasificación

RESUMEN

MSL complexes bind the single male X chromosome in Drosophila to increase transcription approximately 2-fold. Complexes contain at least five proteins and two noncoding RNAs, roX1 and roX2. The mechanism of X chromosome binding is not known. Here, we identify a 110 bp sequence in roX2 characterized by high-affinity MSL binding, male-specific DNase I hypersensitivity, a shared consensus with the otherwise dissimilar roX1 gene, and conservation across species. Mutagenesis of evolutionarily conserved sequences diminishes MSL binding in vivo. MSL binding to these sites is roX RNA dependent, suggesting that complexes become competent for binding only after incorporation of roX RNAs. However, the roX RNA segments homologous to the DNA binding sites are not required, ruling out simple RNA-DNA complementarity as the primary targeting mechanism. Our results are consistent with a model in which nascent roX RNA assembly with MSL proteins is an early step in the initiation of dosage compensation.

Asunto(s)

Compensación de Dosificación (Genética) , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Proteínas Nucleares/metabolismo , Proteínas de Unión al ARN/metabolismo , ARN/metabolismo , Factores de Transcripción/metabolismo , Cromosoma X/genética , Animales , Secuencia de Bases/genética , Sitios de Unión/genética , Desoxirribonucleasa I/genética , Proteínas de Drosophila/genética , Regulación de la Expresión Génica/genética , Marcación de Gen , Datos de Secuencia Molecular , Mutagénesis Sitio-Dirigida/genética , Proteínas Nucleares/genética , ARN/genética , Proteínas de Unión al ARN/genética , Factores de Transcripción/genética , Transcripción Genética/genética