Gene activation in eukaryotes frequently involves interactions between chromosomal regions. We have investigated whether higher-order chromatin structures are involved in the regulation of the Arabidopsis floral repressor gene FLC, a target of several chromatin regulatory pathways. Here, we identify a gene loop involving the physical interaction of the 5 0 and 3 0 flanking regions of the FLC locus using chromosome conformation capture. The FLC loop is unaffected by mutations disrupting conserved chromatin regulatory pathways leading to very different expression states. However, the loop is disrupted during vernalization, the cold-induced, Polycomb-dependent epigenetic silencing of FLC. Loop disruption parallels timing of the cold-induced FLC transcriptional shut-down and upregulation of FLC antisense transcripts, but does not need a cold-induced PHD protein required for the epigenetic silencing. We suggest that gene loop disruption is an early step in the switch from an expressed to a Polycomb-silenced state.
The RNA-binding proteins FCA and FPA were identified based on their repression of the flowering time regulator FLC but have since been shown to have widespread roles in the Arabidopsis thaliana genome. Here, we use whole-genome tiling arrays to show that a wide spectrum of genes and transposable elements are misexpressed in the fca-9 fpa-7 (fcafpa) double mutant at two stages of seedling development. There was a significant bias for misregulated genomic segments mapping to the 3′ region of genes. In addition, the double mutant misexpressed a large number of previously unannotated genomic segments corresponding to intergenic regions. We characterized a subset of these misexpressed unannotated segments and established that they resulted from extensive transcriptional read-through, use of downstream polyadenylation sites, and alternative splicing. In some cases, the transcriptional read-through significantly reduced expression of the associated genes. FCA/FPAdependent changes in DNA methylation were found at several loci, supporting previous associations of FCA/FPA function with chromatin modifications. Our data suggest that FCA and FPA play important roles in the A. thaliana genome in RNA 3′ processing and transcription termination, thus limiting intergenic transcription.
In recent years, photosynthetic microalgae regained attention for biodiesel production. For efficient utilization of microalgae, a number of criteria including a strain with high biomass and lipid productivities and employment of effective and reliable methods for oil extraction from the obtained biomass should be met. Recently, we have isolated and identified three thermo-resistant green microalgae strains, namely; Scenedesmus sp. ME02, Hindakia tetrachotoma ME03 and Micractinium sp. ME05. In this study, we compared percent lipid content of thermos-tolerant microalgal strains using the following solvent extraction methods: Soxhlet, Bligh and Dyer and Folch methods with or without assisted cell disruption techniques including lyophilization, homogenization, ultrasonication, bead and microwave-assisted. The highest increase in lipid yield was obtained with a combination of lyophilization and ultrasonication techniques together with Soxhlet method: 27% of total dry weight for Micractinium sp. ME05. We conclude that lyophilization and ultrasonication are effective assistance methods for lipid extraction from thermo-resistant microalgae.
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