Genetic interactions and dosage effects of Polycomb group genes of Drosophila

The Polycomb (Pc) group of genes are required for maintenance of cell determination in Drosophila melanogaster. At least 11 Pc group genes have been described and there may be up to 40; all are required for normal regulation of homeotic genes, but as a group, their phenotypes are rather diverse. It has been suggested that the products of Pc group genes might be members of a heteromeric complex that acts to regulate the chromatin structure of target loci. We examined the phenotypes of adult flies heterozygous for every pairwise combination of Pc group genes in an attempt to subdivide the Pc group functionally. The results support the idea that Additional sex combs (Asx), Pc, Polycomblike (Pcl), Posterior sex combs (Psc), Sex combs on midleg (Scm), and Sex combs extra (Sce) have similar functions in some imaginal tissues. We show genetic interactions among extra sex combs (esc) and Asx, Enhancer of Pc, Pcl, Enhancer of zeste E(z), and super sex combs and reassess the idea that most Pc group genes function independently of esc. Most duplications of Pc group genes neither exhibit anterior transformations nor suppress the extra sex comb phenotype of Pc group mutations, suggesting that not all Pc group genes behave as predicted by the mass-action model. Surprisingly, duplications of E(z) enhance homeotic phenotypes of esc mutants. Flies with increasing doses of esc ⁺ exhibit anterior transformations, but these are not enhanced by mutations in trithorax group genes. The results are discussed with respect to current models of Pc group function.     

Uncovering a tumor-suppressor function for Drosophila Polycomb group genes

Comment on: Martinez AM, et al. Nat Genet 2009; 41:1076-82.     

Genetic interactions between the Polycomb locus and the Antennapedia and Bithorax complexes of Drosophila

Summary We have studied the embryonic and adult phenotypes of genetic combinations between Polycomb (Pc), Regulator of bithorax (Rg-bx) and the genes of the Bithorax complex (BX-C) and the Antennapedia complex (ANT-C). The products of Pc and Rg-bx genes act antagonistically, their mutant combinations leading to the ectopic expression of genes of the BX-C and ANT-C. The genetic analysis of the Pc locus suggests it is a complex gene. Pc⁺ products behave as members of a regulatory set that negatively control the expression of BX-C and ANT-C genes. Genetic combinations between different doses of Pc, Rg-bx and the genes of the BX-C and ANT-C have phenotypes which may be interpreted as resulting from ectopic derepression of posterior selector genes repressing selector genes of anterior segments. The transformation phenotypes of certain genetic combinations differ in embryos and adults. A model of regulation of the BX-C and the ANT-C genes during the imaginal cell proliferation is presented, in which the specification state is maintained by self-activation of a given selector gene and down modulation of other selector genes in the same cell.     

Maintenance of the engrailed expression pattern by Polycomb group genes in Drosophila.

The stable maintenance of expression patterns of homeotic genes depends on the function of a number of negative trans-regulators, termed the Polycomb (Pc) group of genes. We have examined the pattern of expression of the Drosophila segment polarity gene, engrailed (en), in embryos mutant for several different members of the Pc group. Here we report that embryos mutant for two or more Pc group genes show strong ectopic en expression, while only weak derepression of en occurs in embryos mutant for a single Pc group gene. This derepression is independent of two known activators of en expression: en itself and wingless. Additionally, in contrast to the strong ectopic expression of homeotic genes observed in extra sex combs- (esc-) mutant embryos, the en expression pattern is nearly normal in esc- embryos. This suggests that the esc gene product functions in a pathway independent of the other genes in the group. The data indicate that the same group of genes is required for stable restriction of en expression to a striped pattern and for the restriction of expression of homeotic genes along the anterior-posterior axis, and support a global role for the Pc group genes in stable repression of activity of developmental selector genes.     

Regulation of Polycomb group genes Psc and Su(z)2 in Drosophila melanogaster

Certain Polycomb group (PcG) genes are themselves targets of PcG complexes. Two of these constitute the Drosophila Psc-Su(z)2 locus, a region whose chromatin is enriched for H3K27me3 and contains several putative Polycomb response elements (PREs) that bind PcG proteins. To understand how PcG mechanisms regulate this region, the repressive function of the PcG protein binding sites was analyzed using reporter gene constructs. We find that at least two of these are functional PREs that can silence a reporter gene in a PcG-dependent manner. One of these two can also display anti-silencing activity, dependent on the context. A PcG protein binding site near the Psc promoter behaves not as a silencer but as a down-regulation module that is actually stimulated by the Pc gene product but not by other PcG products. Deletion of one of the PREs increases the expression level of Psc and Su(z)2 by twofold at late embryonic stages. We present evidence suggesting that the Psc-Su(z)2 locus is flanked by insulator elements that may protect neighboring genes from inappropriate silencing. Deletion of one of these regions results in extension of the domain of H3K27me3 into a region containing other genes, whose expression becomes silenced in the early embryo.     

Regulation of cellular plasticity in Drosophila imaginal disc cells by the Polycomb group, trithorax group and lama genes

Drosophila imaginal disc cells can switch fates by transdetermining from one determined state to another. We analyzed the expression profiles of cells induced by ectopic Wingless expression to transdetermine from leg to wing by dissecting transdetermined cells and hybridizing probes generated by linear RNA amplification to DNA microarrays. Changes in expression levels implicated a number of genes: lamina ancestor, CG12534 (a gene orthologous to mouse augmenter of liver regeneration), Notch pathway members, and the Polycomb and trithorax groups of chromatin regulators. Functional tests revealed that transdetermination was significantly affected in mutants for lama and seven different PcG and trxG genes. These results validate our methods for expression profiling as a way to analyze developmental programs, and show that modifications to chromatin structure are key to changes in cell fate. Our findings are likely to be relevant to the mechanisms that lead to disease when homologs of Wingless are expressed at abnormal levels and to the manifestation of pluripotency of stem cells.     

Polycomb group genes are required for neural stem cell survival in postembryonic neurogenesis of Drosophila

Genes of the Polycomb group (PcG) are part of a cellular memory system that maintains appropriate inactive states of Hox gene expression in Drosophila. Here, we investigate the role of PcG genes in postembryonic development of the Drosophila CNS. We use mosaic-based MARCM techniques to analyze the role of these genes in the persistent larval neuroblasts and progeny of the central brain and thoracic ganglia. We find that proliferation in postembryonic neuroblast clones is dramatically reduced in the absence of Polycomb, Sex combs extra, Sex combs on midleg, Enhancer of zeste or Suppressor of zeste 12. The proliferation defects in these PcG mutants are due to the loss of neuroblasts by apoptosis in the mutant clones. Mutation of PcG genes in postembryonic lineages results in the ectopic expression of posterior Hox genes, and experimentally induced misexpression of posterior Hox genes, which in the wild type causes neuroblast death, mimics the PcG loss-of-function phenotype. Significantly, full restoration of wild-type-like properties in the PcG mutant lineages is achieved by blocking apoptosis in the neuroblast clones. These findings indicate that loss of PcG genes leads to aberrant derepression of posterior Hox gene expression in postembryonic neuroblasts, which causes neuroblast death and termination of proliferation in the mutant clones. Our findings demonstrate that PcG genes are essential for normal neuroblast survival in the postembryonic CNS of Drosophila. Moreover, together with data on mammalian PcG genes, they imply that repression of aberrant reactivation of Hox genes may be a general and evolutionarily conserved role for PcG genes in CNS development.     

Genetic interactions of modifier genes and modifiable alleles in Drosophila melanogaster

We have examined the effects of mutations in the six allele-specific modifier genes su(Hw), e(we), su(f), su(s), su(wa), and su(pr) on the expression of 18 modifiable alleles, situated at 11 loci. Ten of the modifiable alleles are associated with insertions of the gypsy retrotransposon and the others include alleles associated with insertions of copia and 412. We tested or retested 90 of the 108 possible combinations and examined the expression of modifiable alleles in flies mutant for pairs of modifier genes in various heterozygous and homozygous configurations. Our principal findings are: (1) a screen of 40,000 mutagenized X chromosomes yielded three new mutations in known modifier genes, but revealed no new modifier genes; (2) the modification effects of different mutations in a given modifier gene were qualitatively similar; (3) each of the six modifiers suppressed some modifiable alleles, enhanced others, and had no noticeable effect on still others; (4) the modifier genes could be placed in four classes, according to their effects on the gypsy-insertion alleles; and (5) the effects of mutations in different modifier genes combined additively. Implications of these results for models of modifier gene action are discussed.     

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.     

Homoeosis in Drosophila. II. a Genetic Analysis of Polycomb

Three dominant mutant alleles of the Polycomb locus of Drosophila melanogaster are associated with homoeotic transformations of meso- and metathoracic to prothoracic legs, a homoeotic transformation of antennae to legs, and abnormalities of wings and some thoracic bristles. Puro and Nygrén (1975) localized Polycomb in the proximal left arm of chromosome 3 within salivary gland chromosome interval 77E,F-80. In the present study, the location and dosage relationships of this locus were examined, using translocation-generated segmental aneuploidy. The results indicate that Polycomb lies within interval 78C,D-79D, and that the locus is haplo-insufficient. Males hypoploid for this interval show meso- and metathoracic leg transformations, and both males and females show wing abnormalities. In addition, the legs of hypoploids of both sexes are shorter than those of wild-type flies, and show aberrancies of segmentation, chaetal number and distribution, and other morphological characteristics. Hypoploid flies do not express a homoeotic antennal-leg transformation, but the deficiency is associated with a Minute phenotype that is known to suppress this transformation in Polycomb flies; thus it cannot be ascertained whether the antennal-leg transformation is a haplo-insufficient phenotype. It is suggested that the expression of non-homoeotic pleiotropic effects provides a criterion for identifying homoeotic mutations that do not function directly in the establishment of determined states, but rather cause homoeosis indirectly. Polycomb is interpreted in this fashion, and it is suggested that the mutant syndrome may result from localized cell death.