Do the BEAF insulator proteins regulate genes involved in cell polarity and neoplastic growth?

AbstractIt was reported that a chromosome with the BEAFNP6377 (NP6377) allele leads to a loss of cell polarity and neoplastic growth in Drosophila melanogaster when homozygous (Gurudatta et al., 2012). We had previously generated the BEAFAB-KO (AB-KO) allele by homologous recombination and did not note these phenotypes (Roy et al., 2007). Both alleles are null mutations. It was unclear why two null alleles of the same gene would give different phenotypes. To resolve this, we performed genetic tests to explore the possibility that the chromosome with the NP6377 allele contained other, second site mutations that might account for the different phenotypes. We found that the chromosome with NP6377 has at least two additional mutations. At least one of these, possibly in combination with the NP6377 allele, is presumably responsible for the reported effects on gene expression, cell polarity and neoplastic growth

Targeted gene replacement by homologous recombination in Drosophila stimulates production of second-site mutations

Gene replacement by homologous recombination is a powerful technique for generating mutations in Drosophila. While using this technique for the BEAF gene, we encountered non-targeted lethal mutations on the target chromosome that complicated the analysis of the BEAF mutations until they were discovered and removed by meiotic recombination. Subsequent experiments indicated that the gene-targeting method leads to a modest but significant three-fold increase in the rate of production of non-targeted lethal mutations. It is important to be aware of this phenomenon when using this method. © 2010 Landes Bioscience

Overlapping but distinct sequences play roles in the insulator and promoter activities of the drosophila BEAF-dependent scs⇔ insulator

Copyright © 2020 by the Genetics Society of America. Chromatin domain insulators are thought to help partition the genome into genetic units called topologically associating domains (TADs). In Drosophila, TADs are often separated by inter-TAD regions containing active housekeeping genes and associated insulator binding proteins. This raises the question of whether insulator binding proteins are involved primarily in chromosomal TAD architecture or gene activation, or if these two activities are linked. The Boundary Element-Associated Factor of 32 kDa (BEAF-32, or BEAF for short) is usually found in inter-TADs. BEAF was discovered based on binding to the scs’ insulator, and is important for the insulator activity of scs’ and other BEAF binding sites. There are divergent promoters in scs’ with a BEAF binding site by each. Here, we dissect the scs’ insulator to identify DNA sequences important for insulator and promoter activity, focusing on the half of scs’ with a high affinity BEAF binding site. We find that the BEAF binding site is important for both insulator and promoter activity, as is another sequence we refer to as LS4. Aside from that, different sequences play roles in insulator and promoter activity. So while there is overlap and BEAF is important for both, insulator and promoter activity can be separated

A genetic screen supports a broad role for the Drosophila insulator proteins BEAF-32A and BEAF-32B in maintaining patterns of gene expression

Chromatin domain insulators are thought to insulate adjacent genes, including their regulatory elements, from each other by organizing chromatin into functionally independent domains. Thus insulators should play a global role in gene regulation by keeping regulatory domains separated. However, this has never been demonstrated. We previously designed and characterized a transgene that is under GAL4 UAS control and encodes a dominant-negative form of the Boundary Element-Associated Factors BEAF-32A and BEAF-32B. The BID transgene encodes the BEAF self-interaction domain but lacks a DNA binding domain. Expression of BID in eye imaginal discs leads to a rough eye phenotype. Here we screen for dominant mutations that modify this eye phenotype. This assay provides evidence for cross-talk between different classes of insulators, and for a broad role of the BEAF proteins in maintaining patterns of gene expression during eye development. Most identified genes encode other insulator binding proteins, transcription factors involved in head development, or general transcription factors. Because it is unlikely that insulator function is limited to eye development, the present results support the hypothesis that insulators play a widespread role in maintaining global transcription programs. © 2006 Springer-Verlag

The drosophila boundary element-associated factors BEAF-32A and BEAF-32B affect chromatin structure

Binding sites for the Drosophila boundary element-associated factors BEAF-32A and -32B are required for the insulator activity of the scs\u27 insulator. BEAF binds to hundreds of sites on polytene chromosomes, indicating that BEAF-utilizing insulators are an important class in Drosophila. To gain insight into the role of BEAF in flies, we designed a transgene encoding a dominant-negative form of BEAF under GAL4 UAS control. This BID protein encompasses the BEAF self-interaction domain. Evidence is provided that BID interacts with BEAF and interferes with scs\u27 insulator activity and that BEAF is the major target of BID in vivo. BID expression during embryogenesis is lethal, implying that BEAF is required during early development. Expression of BID in eye imaginal discs leads to a rough-eye phenotype, and this phenotype is rescued by a third copy of the BEAF gene. Expression of BID in salivary glands leads to a global disruption of polytene chromatin structure, and this disruption is largely rescued by an extra copy of BEAF. BID expression also enhances position-effect variegation (PEV) of the wm4h allele and a yellow transgene inserted into the pericentric heterochromatin of chromosome 2R, while a third copy of the BEAF gene suppresses PEV of both genes. These results support the hypothesis that BEAF-dependent insulators function by affecting chromatin structure or dynamics. Copyright © 2006 by the Genetics Society of America

The effect of processing parameters on particle size in ammonia-induced precipitation of zirconyl chloride under industrially relevant conditions

The effect of pH of precipitation, starting solution concentration, and agitation levels on the particle size of hydrous zirconia precipitates have been investigated. It was found that all three variables affect the particle size of the hydrous zirconia. The smallest particle size is produced by a 0.81 M starting solution, precipitated at pH 12 with a high agitation level. The pH of precipitation was also found to have a significant impact on the type of hydrous zirconia produced. TGA/DTA, micro combustion and TEM / EDS were used to investigate the difference in the powders produced at pH 3 and 12. This work suggests that powders produced at pH 3 will have a structure similar to Zr[OH]4 whilst those at pH 12 are more likely ZrO[OH]2. XRD and micro-combustion suggest that the powders produced at pH 3 retained ammonium chloride whilst those produced at pH 12 did not. The filtration rates for the pH 3 product were significantly faster than that of the powders made at pH 12 which is significant in the industrial production of these materials