Chromatin regulates origin activity in Drosophila follicle cells.

It is widely believed that DNA replication in multicellular animals (metazoa) begins at specific origins to which a pre-replicative complex (pre-RC) binds. Nevertheless, a consensus sequence for origins has yet to be identified in metazoa. Origin identity can change during development, suggesting that there are epigenetic influences. A notable example of developmental specificity occurs in Drosophila, where somatic follicle cells of the ovary transition from genomic replication to exclusive re-replication at origins that control amplification of the eggshell (chorion) protein genes. Here we show that chromatin acetylation is critical for this developmental transition in origin specificity. We find that histones at the active origins are hyperacetylated, coincident with binding of the origin recognition complex (ORC). Mutation of the histone deacetylase (HDAC) Rpd3 induced genome-wide hyperacetylation, genomic replication and a redistribution of the origin-binding protein ORC2 in amplification-stage cells, independent of effects on transcription. Tethering Rpd3 or Polycomb proteins to the origin decreased its activity, whereas tethering the Chameau acetyltransferase increased origin activity. These results suggest that nucleosome acetylation and other epigenetic changes are important modulators of origin activity in metazoa.

Worker education level is a factor in self-compliance with dust-preventive methods among small-scale agate industrial workers.

OBJECTIVES: High incidences of silicosis are continuing to be reported among the agate workers of small-scale household agate processing units in the Khambhat region of Gujarat (India). The objective of this study was to investigate reasons behind the high prevalence of silicosis, and factors affecting the noncompliance with preventive methods among agate workers. METHODS: The study was conducted using a questionnaire-based structured interview method among 82 agate workers in Khambhat to assess their awareness level about silicosis and preventive methods, existing morbidity, worker's attitude toward health, and the prevalence of actual use of preventive methods to avoid silica exposure. RESULTS: The majority of the workers (55%) were aware of silicosis and the harmful effects of silica dust exposure (72%) and knew about simple preventive methods to avoid silica dust exposure (80%), but only a minority of the workers (22%) were actually using the simple and available dust-preventive methods. Only 9% of the uneducated workers were using the preventive methods, while usage was higher among educated workers (28%), who had five or more years of schooling, and these workers had fewer health conditions or less morbidity. Gender and job duration had no effect on the usage of dust-preventive methods. CONCLUSIONS: The data suggest that noncompliance with use of dust-preventive methods could be the reason behind the higher prevalence of silicosis and health morbidity in agate workers, and that years of schooling plays a significant role in the increased usage and self-compliance with dust-preventive methods among agate workers.

Drosophila double-parked is sufficient to induce re-replication during development and is regulated by cyclin E/CDK2.

It is important that chromosomes are duplicated only once per cell cycle. Over-replication is prevented by multiple mechanisms that block the reformation of a pre-replicative complex (pre-RC) onto origins in S and G2 phase. We have investigated the developmental regulation of Double-parked (Dup) protein, the Drosophila ortholog of Cdt1, a conserved and essential pre-RC component found in human and other organisms. We find that phosphorylation and degradation of Dup protein at G1/S requires cyclin E/CDK2. The N terminus of Dup, which contains ten potential CDK phosphorylation sites, is necessary and sufficient for Dup degradation during S phase of mitotic cycles and endocycles. Mutation of these ten phosphorylation sites, however, only partially stabilizes the protein, suggesting that multiple mechanisms ensure Dup degradation. This regulation is important because increased Dup protein is sufficient to induce profound rereplication and death of developing cells. Mis-expression has different effects on genomic replication than on developmental amplification from chorion origins. The C terminus alone has no effect on genomic replication, but it is better than full-length protein at stimulating amplification. Mutation of the Dup CDK sites increases genomic re-replication, but is dominant negative for amplification. These two results suggest that phosphorylation regulates Dup activity differently during these developmentally specific types of DNA replication. Moreover, the ability of the CDK site mutant to rapidly inhibit BrdU incorporation suggests that Dup is required for fork elongation during amplification. In the context of findings from human and other cells, our results indicate that stringent regulation of Dup protein is critical to protect genome integrity.