A Polycomb response element in the Ubx gene that determines an epigenetically inherited state of repression.

Segmentation genes provide the signals for the activation and regulation of homeotic genes in Drosophila but cannot maintain the resulting pattern of expression because their activity ceases halfway through embryogenesis. Maintenance of the pattern is due to the Polycomb group of genes (Pc-G) and the trithorax group of genes (trx-G), responsible for the persistence of the active or repressed state of homeotic genes. We have identified a regulatory element in the Ubx gene that responds to Pc-G and trx-G genes. Transposons carrying this element create new binding sites for Pc-G products in the polytene chromosomes. This Pc-G maintenance element (PRE), establishes a repressive complex that keeps enhancers repressed in cells in which they were originally repressed and maintains this state through many cell divisions. The trx-G products stimulate the expression of enhancers in cells in which they were originally active. This mechanism is responsible for the correct regulation of imaginal disc enhancers, which lack themselves antero-posterior positional information. The PRE also causes severe variegation of the mini-white gene present in the transposon, a phenomenon very similar to heterochromatic position-effect variegation. The significance of this mechanism for homeotic gene regulation is discussed.     

DSP1, an HMG-like protein, is involved in the regulation of homeotic genes

The Drosophila dsp1 gene, which encodes an HMG-like protein, was originally identified in a screen for corepressors of Dorsal. Here we report that loss of dsp1 function causes homeotic transformations resembling those associated with loss of function in the homeotic genes Sex combs reduced (Scr), Ultrabithorax (Ubx), and Abdominal-B. The expression pattern of Scr is altered in dsp1 mutant imaginal discs, indicating that dsp1 is required for normal expression of this gene. Genetic interaction studies reveal that a null allele of dsp1 enhances trithorax-group gene (trx-G) mutations and partially suppresses Polycomb-group gene (Pc-G) mutations. On the contrary, overexpression of dsp1 induces an enhancement of the transformation of wings into halteres and of the extra sex comb phenotype of Pc. In addition, dsp1 male mutants exhibit a mild transformation of A4 into A5. Comparison of the chromatin structure at the Mcp locus in wild-type and dsp1 mutant embryos reveals that the 300-bp DNase I hypersensitive region is absent in a dsp1 mutant context. We propose that DSP1 protein is a chromatin remodeling factor, acting as a trx-G or a Pc-G protein depending on the considered function.     

corto genetically interacts with Pc-G and trx-G genes and maintains the anterior boundary of Ultrabithorax expression in Drosophila larvae

In Drosophila melanogaster, segment identity is determined by specific expression of homeotic genes (Hox). The Hox expression pattern is first initiated by gap and pair-rule genes and then maintained by genes of the Polycomb-group (Pc-G) and the trithorax-group (trx-G). The corto gene is a putative regulator of the Hox genes since mutants exhibit homeotic transformations. We show here that, in addition to previously reported genetic interactions with the Pc-G genes Enhancer of zeste, Polycomb and polyhomeotic, mutations in corto enhance the extra-sex-comb phenotype of multi sex combs, Polycomb-like and Sex combs on midleg. corto also genetically interacts with a number of trx-G genes (ash1, kismet, kohtalo, moira, osa, Trithorax-like and Vha55). The interactions with genes of the trx-G lead to phenotypes displayed in the wing, in the postpronotum or in the thoracic mechanosensory bristles. In addition, we analyzed the regulation of the Hox gene Ultrabithorax (Ubx) in corto mutants. Our results provide evidence that corto maintains the anterior border of Ubx expression in third-instar larvae. We suggest that this regulation is accomplished through an interaction with the products of the Pc-G and trx-G genes.     

Dominant modifiers of the polyhomeotic extra-sex-combs phenotype induced by marked P element insertional mutagenesis in Drosophila.

Members of the Polycomb group (Pc-G) and trithorax group (trx-G) of genes, as well as the enhancers of trx-G and Pc-G (ETP), function together to maintain segment identity during Drosophila development. In order to obtain new marked P mutations in these genes, we screened for dominant modifiers of the extra-sex-combs phenotype displayed by males mutant for the polyhomeotic (ph) gene, a member of the Pc-G group. Five P(lacW) insertions in four different genes were found to stably suppress ph: two are allelic to trithorax, one is the first allele specific to the Minute(2)21C gene, and the remaining two define new trx-G genes, toutatis (tou) in 48A and taranis (tara) in 89B10-13. tou is predicted to encode a 3109 amino acid sequence protein (TOU), which contains a TAM DNA-binding domain, a WAKZ motif, two PHD zinc fingers and a C-terminal bromodomain, and as such is likely to be involved in regulation of chromatin structure as a subunit of a novel chromatin remodelling complex. In a previous study, we found that insertion of a P(ph) transposable element containing ph regulatory sequences creates a high frequency of mutations modifying ph homeotic phenotypes. One such insertion enhanced the ph phenotype and we show that it is a new allele of UbcD1/eff, a gene encoding a ubiquitin-conjugating enzyme that is involved in telomere association and potentially in chromatin remodelling.     

DSP1, a Drosophila HMG protein, is involved in spatiotemporal expression of the homoeotic gene Sex combs reduced

BACKGROUND INFORMATION: The Pc-G (Polycomb group) and trx-G (trithorax group) genes play a key role in the regulation of the homoeotic genes. The homoeotic gene Scr (Sex combs reduced) contained in the Antennapedia complex specifies segmental identity of the labial and prothoracic segments in Drosophila. Regulation of Scr requires the action of different enhancer elements spread over several kilobases. We previously identified an HMGB (high mobility group)-like protein DSP1 (dorsal switch protein 1), which works like a trx-G protein for the normal Scr expression. RESULTS: In the present study, we attempted to characterize the regulatory sequences involved in the maintenance of the Scr activation by DSP1. We report here, using a transgenic line for the Scr10.0XbaI-regulatory element, that lack of DSP1 affects the function of a reporter gene in legs' imaginal discs but not in embryos. We show by immunolocalization that DSP1 is recruited on polytene chromosomes to the insertion site of the transgene. Moreover, using chromatin immunoprecipitation experiments, we identify two regions of 1 kb in Scr10.0XbaI as the main DSP1 targets. CONCLUSION: These results provide strong evidence that the Scr gene expression is influenced by direct interaction between DSP1 and two Scr regulation elements. In addition, our results show that this interaction undergoes dynamic changes during development.     

The bromodomain protein LIN-49 and trithorax-related protein LIN-59 affect development and gene expression in Caenorhabditis elegans

We have molecularly characterized the lin-49 and lin-59 genes in C. elegans, and found their products are related to Drosophila trithorax group (trx-G) proteins and other proteins implicated in chromatin remodelling. LIN-49 is structurally most similar to the human bromodomain protein BR140, and LIN-59 is most similar to the Drosophila trx-G protein ASH1. In C. elegans, lin-49 and lin-59 are required for the normal development of the mating structures of the adult male tail, for the normal morphology and function of hindgut (rectum) cells in both males and hermaphrodites and for the maintenance of structural integrity in the hindgut and egg-laying system in adults. Expression of the Hox genes egl-5 and mab-5 is reduced in lin-49 and lin-59 mutants, suggesting lin-49 and lin-59 regulate HOM-C gene expression in C. elegans as the trx-G genes do in Drosophila. lin-49 and lin-59 transgenes are expressed widely throughout C. elegans animals. Thus, in contrast to the C. elegans Polycomb group (Pc-G)-related genes mes-2 and mes-6 that function primarily in the germline, we propose lin-49 and lin-59 function in somatic development similar to the Drosophila trx-G genes.     

Genomic imprinting in Drosophila is maintained by the products of Suppressor of variegation and trithorax group,but not Polycomb group,genes

Genomic imprinting is a form of epigenetic inheritance that is characterized by differential expression of a gene depending on its parental origin. The mini-X chromosome Dp(1;f)LJ9 in Drosophila shows this type of classical imprinting; when transmitted by the maternal parent genes on this chromosome are fully expressed, but when the chromosome is transmitted by the male parent at least three genes are subject to silencing, resulting in a variegated expression pattern. Chemical and environmental modifiers of position-effect variegation have been shown to alter the somatic maintenance of the imprint. To extend these observations, several mutations in chromatin-associated proteins were examined for their effect on imprinting on the Dp(1;f)LJ9 mini-X chromosome. Effects on establishment and maintenance were independently assessed by genetically associating the mutations in chromatin modifiers with the mini-X chromosome in either the parents, where the imprint is established, or the progeny, in which the imprint must be maintained. Nine Suppressor of variegation [ Su(var)] mutations, including alleles of the Su(var)2-5 gene, which encodes the well characterized heterochromatin-associated protein HP1, abolished maintenance but not the establishment of the imprint. Mutant alleles of two genes in the trithorax group ( trx-G), brahma and trithorax, showed a maternal-effect enhancement of the paternal imprint. Surprisingly, however, with the exception of an Enhancer of Polycomb [ E(Pc)] allele, none of the Polycomb-group ( Pc-G) mutations tested affected the imprint. Thus, the maintenance of this imprint relies on the wild-type products of Su(var) and trx-G, but not Pc-G, genes. Finally, none of the mutations tested affected the maintenance of the maternal imprint or the establishment of either the maternal or paternal imprint, suggesting that the maternal and paternal imprints depend on different molecular processes and that imprint establishment and maintenance are independently regulated.     

Genetic factors controlling white gene expression of the transposon A(R) 4-24 at a telomere in Drosophila melanogaster.

The position effect of the AR 4-24 P[white, rosy] transposon was studied at cytological position 60F. Three copies of the transposon (within approximately 50-kb region) resulted in a spatially restricted pattern of white variegation. This pattern was modified by temperature and by removal of the Y chromosome, suggesting that it was due to classical heterochromatin-induced position effect variegation (PEV). In contrast with classical PEV, extra dose of the heterochromatin protein 1 (HP1) suppressed white variegation and one dose enhanced it. The effect of Pc-G, trx-G, and other PEV suppressors was also tested. It was found that E(Pc)1, TrlR85, and mutations of Su(z)2C relieve A(R) 4-24-silencing and z1 enhances it. To explain the results obtained with these modifiers, it is proposed that PEV and telomeric position effect can counteract each other at this particular cytological site.     

The Iroquois homeobox genes function as dorsal selectors in the Drosophila head

The Iroquois complex (Iro-C) genes are expressed in the dorsal compartment of the Drosophila eye/antenna imaginal disc. Previous work has shown that the Iro-C homeoproteins are essential for establishing a dorsoventral pattern organizing center necessary for eye development. Here we show that, in addition, the Iro-C products are required for the specification of dorsal head structures. In mosaic animals, the removal of the Iro-C transforms the dorsal head capsule into ventral structures, namely, ptilinum, prefrons and suborbital bristles. Moreover, the Iro-C(-) cells can give rise to an ectopic antenna and maxillary palpus, the main derivatives of the antenna part of the imaginal disc. These transformations are cell-autonomous, which indicates that the descendants of a dorsal Iro-C(-) cell can give rise to essentially all the ventral derivatives of the eye/antenna disc. These results support a role of the Iro-C as a dorsal selector in the eye and head capsule. Moreover, they reinforce the idea that developmental cues inherited from the distinct embryonic segments from which the eye/antenna disc originates play a minimal role in the patterning of this disc.     

A Role of Polycomb Group Genes in the Regulation of Gap Gene Expression in Drosophila

Anteroposterior polarity of the Drosophila embryo is initiated by the localized activities of the maternal genes, bicoid and nanos, which establish a gradient of the hunchback (hb) morphogen. nanos determines the distribution of the maternal Hb protein by regulating its translation. To identify further components of this pathway we isolated suppressors of nanos. In the absence of nanos high levels of Hb protein repress the abdomen-specific genes knirps and giant. In suppressor-of-nanos mutants, knirps and giant are expressed in spite of high Hb levels. The suppressors are alleles of Enhancer of zeste (E(z)) a member of the Polycomb group (Pc-G) of genes. We show that E(z), and likely other Pc-G genes, are required for maintaining the expression domains of knirps and giant initiated by the maternal Hb protein gradient. We have identified a small region of the knirps promoter that mediates the regulation by E(z) and hb. Because Pc-G genes are thought to control gene expression by regulating chromatin, we propose that imprinting at the chromatin level underlies the determination of anteroposterior polarity in the early embryo.     

Dorso-ventral asymmetric functions of teashirt in Drosophila eye development depend on spatial cues provided by early DV patterning genes

The teashirt (tsh) gene has dorso-ventral (DV) asymmetric functions in Drosophila eye development: promoting eye development in dorsal and suppressing eye development in ventral by Wingless mediated Homothorax (HTH) induction [Development 129 (2002) 4271]. We looked for DV spatial cues required by tsh for its asymmetric functions. The dorsal Iroquois-Complex (Iro-C) genes and Delta (Dl) are required and sufficient for the tsh dorsal functions. The ventral Serrate (Ser), but not fringe (fng) or Lobe (L), is required and sufficient for the tsh ventral function. We propose that DV asymmetric function of tsh represents a novel tier of DV pattern regulation, which takes place after the spatial expression patterns of early DV patterning genes are established in the eye.     

Histone acetylation and hSWI/SNF remodeling act in concert to stimulate V(D)J cleavage of nucleosomal DNA

The ordered assembly of immunoglobulin and TCR genes by V(D)J recombination depends on the regulated accessibility of individual loci. We show here that the histone tails and intrinsic nucleosome structure pose significant impediments to V(D)J cleavage. However, alterations to nucleosome structure via histone acetylation or by stable hSWI/SNF-dependent remodeling greatly increase the accessibility of nucleosomal DNA to V(D)J cleavage. Moreover, acetylation and hSWI/SNF remodeling can act in concert on an individual nucleosome to achieve levels of V(D)J cleavage approaching those observed on naked DNA. These results are consistent with a model in which regulated recruitment of chromatin modifying activities is involved in mediating the lineage and stage-specific control of V(D)J recombination.