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The complexity of regulatory systems in higher eukaryotes, featuring many distantly located enhancers that nonetheless properly activate the target promoters, has prompted the hypothesis that the action of enhancers should be restricted by insulators. Continuing our research on the functional role of insulators and the consequences of their interaction in Drosophila, we studied the interplay of different Su(Hw)-dependent Drosophila insulators. The set of transgenic constructs comprised two consecutive genes (yellow and white) with their enhancers and insulator elements differently arranged in between and/or around the gene(s). All insulators were found to interact in twin or mixed tandems, demonstrating the bypass phenomenon. However, insulator pairing around a gene did not always improve its isolation from an outside enhancer. On the other hand, merely two insulator elements (identical or different) in appropriate positions can permit the expression of one gene but not the gene next to it or, conversely, largely block the transcription of the first gene, while allowing full enhancement of the second, or make them behave similarly. Thus, the results of this study support the model that loop formation by insulators is an essential component of insulator action on a positive and negative regulation of an enhancer-promoter communication.

Much of the research on insulators in Drosophila has been done with transgenic constructs using the white gene (mini-white) as reporter. Hereby we report that the sequence between the white and CG32795 genes in Drosophila melanogaster contains an insulator of a novel kind. Its functional core is within a 368 bp segment almost contiguous to the white 3'UTR, hence we name it as Wari (white-abutting resident insulator). Though Wari contains no binding sites for known insulator proteins and does not require Su(Hw) or Mod(mdg4) for its activity, it can equally well interact with another copy of Wari and with unrelated Su(Hw)-dependent insulators, gypsy or 1A2. In its natural downstream position, Wari reinforces enhancer blocking by any of the three insulators placed between the enhancer and the promoter; again, Wari-Wari, Wari-gypsy or 1A2-Wari pairing results in mutual neutralization (insulator bypass) when they precede the promoter. The distressing issue is that this element hides in all mini-white constructs employed worldwide to study various insulators and other regulatory elements as well as long-range genomic interactions, and its versatile effects could have seriously influenced the results and conclusions of many works.

Chromatin insulators affect interactions between promoters and enhancers/silencers and function as barriers for spreading of repressive chromatin. The Su(Hw) protein is responsible for activity of the best-studied Drosophila insulators. Here we demonstrate that an evolutionarily conserved protein, E(y)2/Sus1, is recruited to the Su(Hw) insulators via binding to the zinc-finger domain of Su(Hw). Partial inactivation of E(y)2 in a weak mutation, e(y)2(u1), impairs only the barrier, but not the enhancer-blocking, activity of the Su(Hw) insulators. Whereas neither su(Hw)(-) nor e(y)2(u1) affects fly viability, their combination proves lethal, testifying to functional interaction between Su(Hw) and E(y)2 in vivo. Apparently, different domains of Su(Hw) recruit proteins responsible for enhancer-blocking and for the barrier activity.

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.

One of the dogmas of transcriptional regulation in higher eukaryotes suggests the existence of transcriptional domains with no promoter-enhancer interactions between them. Specific regulatory elements, known as insulators, block the interaction between an enhancer and a promoter. Insulators are believed to act as transcription domain boundaries. The review considers general properties of well-known insulators identified in Drosophila and vertebrates. The mechanism of action of insulators and their role in the regulation of gene expression are discussed on the basis of available information.

Having evaluated work conditions in sugar-beet seeds treatment with furadan, the authors justified use of an individual protective means for the workers. The recommendations are coveralls of nonwoven materials like Tyvek F, C that are resistant against concentrated and working solutions of furadan. To protect respiratory organs against furadan vapors and aerosols, one should use respirator of sorption filter half mask.

Different colourimetric methods for measuring carbohydrates in yeast have been compared. A method using 5% phenol aqueous solution in the presence of concentrated sulphuric acid has been developed to quantitate carbohydrates. The method has been described as applied to an analysis of dry yeast.