O. B. Simonova

A drosophila protein that imparts directionality on a chromatin insulator is an enhancer of position-effect variegation

The suppressor of Hairy wing (su(Hw)) protein inhibits the function of transcriptional enhancers located distally from the promoter with respect to the location of su(Hw)-binding sites. This polarity is due to the ability of the su(Hw)-binding region to form a chromatin insulator. Mutations in modifier of mdg4 (mod(mdg4)) enhance the effect of su(Hw) by inhibiting the function of enhancers located on both sides of the su(Hw)-binding region. This inhibition results in a variegated expression pattern, and mutations in mod(mdg4) act as classical enhancers of position-effect variegation. The mod(mdg4) and su(Hw) proteins interact with each other. The mod(mdg4) protein controls the nature of the repressive effect of su(Hw): in the absence of mod(mdg4) protein, su(Hw) exerts a bidirectional silencing effect, whereas in the presence of mod(mdg4), the silencing effect is transformed into unidirectional repression.

Two Isoforms of Drosophila TRF2 Are Involved in Embryonic Development, Premeiotic Chromatin Condensation, and Proper Differentiation of Germ Cells of Both Sexes

ABSTRACT The Drosophila TATA box-binding protein (TBP)-related factor 2 (TRF2 or TLF) was shown to control a subset of genes different from that controlled by TBP. Here, we have investigated the structure and functions of the trf2 gene. We demonstrate that it encodes two protein isoforms: the previously described 75-kDa TRF2 and a newly identified 175-kDa version in which the same sequence is preceded by a long N-terminal domain with coiled-coil motifs. Chromatography of Drosophila embryo extracts revealed that the long TRF2 is part of a multiprotein complex also containing ISWI. Both TRF2 forms are detected at the same sites on polytene chromosomes and have the same expression patterns, suggesting that they fulfill similar functions. A study of the manifestations of the trf2 mutation suggests an essential role of TRF2 during embryonic Drosophila development. The trf2 gene is strongly expressed in germ line cells of adult flies. High levels of TRF2 are found in nuclei of primary spermatocytes and trophocytes with intense transcription. In ovaries, TRF2 is present both in actively transcribing nurse cells and in the transcriptionally inactive oocyte nuclei. Moreover, TRF2 is essential for premeiotic chromatin condensation and proper differentiation of germ cells of both sexes.

Cerd4, third member of the d4 gene family: expression and organization of genomic locus

Abstract. Two members of the d4 family of presumptive transcription modulators, neuro-d4 (Neud4) and ubi-d4/Requiem (Req), have been characterized previously. We cloned and characterized the third member of this gene family, cer-d4 (Cerd4), from chicken and mouse cDNA libraries. The expression patterns of Cerd4 gene in both species are similar and more restricted than expression patterns of other two d4 genes. The main sites of Cerd4 expression are retina and cerebellum, where multiple transcripts could be detected. Two major types of Cerd4 proteins are a full-length isoform possessing all domains characteristic to the d4 family and truncated XZ isoform without C-terminal tandem of PHD fingers. The developmental kinetics of expression of these isoforms is different. The intron/exon structure of human Cerd4 gene is similar to that of neuro-d4 and ubi-d4/Requiem genes, but most introns of Cerd4 gene are much larger than the corresponding introns of the other two genes.

Expression pattern of dd4, a sole member of the d4 family of transcription factors in Drosophila melanogaster.

In vertebrates, three members of the d4 gene family code for proteins, which are believed to function as transcription factors and involved in regulation of various intracellular processes. One member of the family, ubi-d4/requiem is ubiquitously expressed gene and two other, neuro-d4 and cer-d4, are expressed predominantly in the neural tissues (Nucleic Acids Res. 20 (1992) 5579; Biochim. Biophys. Acta 14 (1992) 172; Mamm. Genome 11 (2000) 72; Mamm. Genome 12 (2001) 862). Typically, d4 proteins show distinct domain organisation with domain 2/3 in the N-terminal, Krüppel-type zinc finger in the central and two adjacent PHD-fingers (d4-domain) in the C-terminal part of the molecule. However, alternative splicing, which is responsible for complex expression patterns of both neurospecific members of the family, generates multiple protein isoforms lacking certain domains (Nucleic Acids Res. 20 (1992) 5579; Genomics 36 (1996) 174; Mamm. Genome 11 (2000) 72; Mamm. Genome 12 (2001) 862). Exact function of d4 proteins is unclear but their involvement in regulation of differentiation and apoptotic cell death has been proposed (J. Biol. Chem. 269 (1994) 29515; Mamm. Genome 11 (2000) 72; Mamm. Genome 12 (2001) 862). Here we identified a single gene, dd4, in the genome of Drosophila melanogaster, the protein product of which could be assigned to the d4 family. Expression of dd4 is regulated during Drosophila development, and is most prominent in syncytial embryos and later in the embryonic nervous and reproductive systems. In flies dd4 mRNA is found in most tissues but the highest level of expression is detected in ovaries.

Combination of Hypomorphic Mutations of the Drosophila Homologues of Aryl Hydrocarbon Receptor and Nucleosome Assembly Protein Family Genes Disrupts Morphogenesis, Memory and Detoxification

Aryl hydrocarbon receptor is essential for biological responses to endogenous and exogenous toxins in mammals. Its Drosophila homolog spineless plays an important role in fly morphogenesis. We have previously shown that during morphogenesis spineless genetically interacts with CG5017 gene, which encodes a nucleosome assembly factor and may affect cognitive function of the fly. We now demonstrate synergistic interactions of spineless and CG5017 in pathways controlling oxidative stress response and long-term memory formation in Drosophila melanogaster. Oxidative stress was induced by low doses of X-ray irradiation of flies carrying hypomorphic mutation of spineless, mutation of CG5017, and their combination. To determine the sensitivity of these mutants to pharmacological modifiers of the irradiation effect, we irradiated flies growing on standard medium supplemented by radiosensitizer furazidin and radioprotector serotonin. The effects of irradiation were investigated by analyzing leg and antenna morphological structures and by using real-time PCR to measure mRNA expression levels for spineless, Cyp6g1 and Gst-theta genes. We also examined long-term memory in these mutants using conditioned courtship suppression paradigm. Our results show that the interaction of spineless and CG5017 is important for regulation of morphogenesis, long-term memory formation, and detoxification during oxidative stress. Since spineless and CG5017 are evolutionary conserved, these results must be considered when evaluating the risk of combining similar mutations in other organisms, including humans.

Source of asymmetry in ontogeny: Early polarization of the germline cyst and oocyte in Drosophila

One or more body axes are already formed in the egg in many vertebrates and invertebrates. In Drosophila, the anterior-posterior and dorsoventral axes are determined during oogenesis owing to the asymmetric localization of the bicoid, oskar, and gurken mRNAs in the oocyte (prospective egg). The localization of these transcripts depends on the polarized organization of the oocyte cytoskeleton and, consequently, the oocyte polarity. Initial asymmetry, leading to the oocyte polarity, is established in early ontogeny, during oocyte determination. The review considers the steps of early polarization and oocyte differentiation in Drosophila, the genetic control of these processes, and the findings that suggest an early oocyte polarity for vertebrates.

P-Ph-Mediated Repression of the leg–arista–wing complexGene Transcription in Drosophila

Preliminary analysis of the leg–arista–wing complex (lawc) gene region in the corresponding mutants revealed Pelement insertion in the transcribed region of this locus. To demonstrate the main role of P element insertion in the complex pleiotropic phenotypic effect of the lawc gene, a system using P-Ph chimeric protein and based on the P-mediated repression of the lawc in vivo transcription was applied. As a result, extreme lawc-mutant phenotypes were obtained and examined. The P-Ph-mediated decrease of the level of the lawc gene transcription was also demonstrated.

Screening of genomic regions affecting lawc/Trf2 gene expression during Drosophila melanogaster development

Abstractleg-arista-wing complex (lawc) mutations affect the expression of Drosophila proteins homologous to a human basic transcription factor, TBP (TATA-box binding protein) Related Factor 2 (TRF2), specifically involved in development.The paper for the first time reports the application of genetic screens for various genomic regions to identify genetic interactions between the lawc/Trf2 gene and other genes and genetic loci by using Deficiency Kit lines with small deletions in total providing maximal coverage of the genome. The deletion mapping allowed us to recover 26 genomic regions that, when deleted, are lethal or modify the mutant phenotype due to a decreased TRF2 expression level. These deletions could be useful in identifying both novel TRF2 targets and its positive and negative regulators.There is evidence that TRF2 can be a component of high molecular DNA Replication-related Element Factor (DREF) and Nucleosome Remodeling Factor (NURF)-containing complexes. The present study for the first time reports new genetic interactions of lawc/Trf2 with genes that encode basic and specific transcription factors. In most cases, if mutated, those genes caused developmental defects or death of progeny. However, in the case of the e(y)1 gene, coding for the Taf9 transcription factor, only the male reproductive system is impaired when the lawc/Trf2 phenotype is associated with a e(y)1 gene mutation. Mutant lawcp1e(y)1u1 males become infertile due to primary spermatocyte maturation arrest and impaired premeiotic chromosome condensation in germ cells.

Overlapping genes and antisense transcription in eukaryotes

Numerous studies showed that overlapping genes are fairly common elements of genome organization, not only in viruses and prokaryotes but also in eukaryotes. At the same time, the regulatory mechanisms of overlapping gene expression, as well as the functional relevance of antisense transcription, are still relatively unknown. This review describes the history of the discovery of regulatory antisense RNAs, the types of gene overlap, and the putative mechanisms of their functioning. In conclusion, the critical views of different authors on the problem of detecting overlapping genes and an evaluation of the level of antisense transcription are presented.

Effect of mutations in lawc/Trf2 gene on chromocenter formation and chromosome segregation in Drosophila melanogaster

In Drosophila, one of the genes of the lawc/Trf2 (leg-arista-wing complex/TBP-related factor 2) complex encodes an alternative basic transcription factor homologous to the TRF2 protein in vertebrates and human and belongs to a conservative Tbp (TATA box-binding protein) gene family. In the present study, reasons for the high frequency of chromosome nondisjunction were studied among descendants of mutants from 18 lines with a decreased expression of the TRF2 protein. It was determined that the suppression of the TRF2 expression violates the development of a compact chromocenter and the correct approach of homologous chromosomes (in germinative and somatic cells). The possibility of the participation of TRF2 in the evolutionary genetically programmed process of sex-ratio meiotic drive, which is typical of a number of animal species, is discussed.