ABSTRACT: In this work, we have studied four closely related members of the five known Drosophila hnRNP A/B family, hrp36, hrp38, hrp40, and hrp48. Using splicing-sensitive microarrays, between 200 and 300 genes were detected as specifically regulated by each individual hrp protein following down regulation of the protein expression level by RNAi-mediated knock-down. Keywords: Genetic modification
Project description:In this work, we have studied four closely related members of the five known Drosophila hnRNP A/B family, hrp36, hrp38, hrp40, and hrp48. Using splicing-sensitive microarrays, between 200 and 300 genes were detected as specifically regulated by each individual hrp protein following down regulation of the protein expression level by RNAi-mediated knock-down. Keywords: Genetic modification
Project description:In this work, we have studied four closely related members of the five known Drosophila hnRNP A/B family, hrp36, hrp38, hrp40, and hrp48. Using splicing-sensitive microarrays, between 200 and 300 genes were detected as specifically regulated by each individual hrp protein following down regulation of the protein expression level by RNAi-mediated knock-down. Keywords: Genetic modification
Project description:In this work, we have studied four closely related members of the five known Drosophila hnRNP A/B family, hrp36, hrp38, hrp40, and hrp48. Using splicing-sensitive microarrays, between 200 and 300 genes were detected as specifically regulated by each individual hrp protein following down regulation of the protein expression level by RNAi-mediated knock-down. Keywords: Genetic modification
Project description:In this work, we have studied four closely related members of the five known Drosophila hnRNP A/B family, hrp36, hrp38, hrp40, and hrp48. Using splicing-sensitive microarrays, between 200 and 300 genes were detected as specifically regulated by each individual hrp protein following down regulation of the protein expression level by RNAi-mediated knock-down. Keywords: Genetic modification This set of arrays evaluate the off target effect of the RNAi. Two differerent non-overlapping dsRNA targeting hrp36 were used to knock down its expression. The net-expression value of each splicing junction was then extracted and compare.
Project description:In this work, we have studied four closely related members of the five known Drosophila hnRNP A/B family, hrp36, hrp38, hrp40, and hrp48. Using splicing-sensitive microarrays, between 200 and 300 genes were detected as specifically regulated by each individual hrp protein following down regulation of the protein expression level by RNAi-mediated knock-down. Keywords: Genetic modification Each individual protein (four total) were individually knock-down. RNA from biological triplicates were isolated 4 days after the initial treatment. A non-specific RNAi control was performed and the RNA isolated from multiple replicates were pooled and used as control sample in the microarray.
Project description:Chromosomal RNAs (cRNAs) are a poorly understood fraction of cellular RNAs that co-purify with chromatin. Here we show that, in Drosophila, cRNAs constitute a heterogeneous group of RNA species that cover ~28% of the genome. Intriguingly, we found that cRNAs are highly enriched in heterochromatic transcripts. Our results show that heterochromatic cRNAs interact with the hnRNP A/B proteins hrp36 and hrp48 to assemble into RNP particles. We also show that depletion of linker histone dH1, a major component of chromatin, impairs assembly of hrp36 and hrp48 onto heterochromatic cRNAs. Concomitantly, impaired cRNAs assembly induces the accumulation of heterochromatic cRNAs and the formation of unscheduled RNA::DNA hybrids (R-loops). Linker histones H1 are known to regulate chromatin structure and compaction and, indeed, we show that dH1 depletion perturbs chromatin organization, reducing nucleosome occupancy and specifically increasing accessibility and 3D interactions within heterochromatin. These perturbations facilitate annealing of cRNAs to the DNA template, enhancing R-loops formation and cRNAs retention at heterochromatin. Altogether, these results unveil the unexpected contribution of linker histones to RNPs assembly and homeostasis of cRNAs.
Project description:Chromosomal RNAs (cRNAs) are a poorly understood fraction of cellular RNAs that co-purify with chromatin. Here we show that, in Drosophila, cRNAs constitute a heterogeneous group of RNA species that cover ~28% of the genome. Intriguingly, we found that cRNAs are highly enriched in heterochromatic transcripts. Our results show that heterochromatic cRNAs interact with the hnRNP A/B proteins hrp36 and hrp48 to assemble into RNP particles. We also show that depletion of linker histone dH1, a major component of chromatin, impairs assembly of hrp36 and hrp48 onto heterochromatic cRNAs. Concomitantly, impaired cRNAs assembly induces the accumulation of heterochromatic cRNAs and the formation of unscheduled RNA::DNA hybrids (R-loops). Linker histones H1 are known to regulate chromatin structure and compaction and, indeed, we show that dH1 depletion perturbs chromatin organization, reducing nucleosome occupancy and specifically increasing accessibility and 3D interactions within heterochromatin. These perturbations facilitate annealing of cRNAs to the DNA template, enhancing R-loops formation and cRNAs retention at heterochromatin. Altogether, these results unveil the unexpected contribution of linker histones to RNPs assembly and homeostasis of cRNAs.
Project description:Chromosomal RNAs (cRNAs) are a poorly understood fraction of cellular RNAs that co-purify with chromatin. Here we show that, in Drosophila, cRNAs constitute a heterogeneous group of RNA species that cover ~28% of the genome. Intriguingly, we found that cRNAs are highly enriched in heterochromatic transcripts. Our results show that heterochromatic cRNAs interact with the hnRNP A/B proteins hrp36 and hrp48 to assemble into RNP particles. We also show that depletion of linker histone dH1, a major component of chromatin, impairs assembly of hrp36 and hrp48 onto heterochromatic cRNAs. Concomitantly, impaired cRNAs assembly induces the accumulation of heterochromatic cRNAs and the formation of unscheduled RNA::DNA hybrids (R-loops). Linker histones H1 are known to regulate chromatin structure and compaction and, indeed, we show that dH1 depletion perturbs chromatin organization, reducing nucleosome occupancy and specifically increasing accessibility and 3D interactions within heterochromatin. These perturbations facilitate annealing of cRNAs to the DNA template, enhancing R-loops formation and cRNAs retention at heterochromatin. Altogether, these results unveil the unexpected contribution of linker histones to RNPs assembly and homeostasis of cRNAs.
Project description:Chromosomal RNAs (cRNAs) are a poorly understood fraction of cellular RNAs that co-purify with chromatin. Here we show that, in Drosophila, cRNAs constitute a heterogeneous group of RNA species that cover ~28% of the genome. Intriguingly, we found that cRNAs are highly enriched in heterochromatic transcripts. Our results show that heterochromatic cRNAs interact with the hnRNP A/B proteins hrp36 and hrp48 to assemble into RNP particles. We also show that depletion of linker histone dH1, a major component of chromatin, impairs assembly of hrp36 and hrp48 onto heterochromatic cRNAs. Concomitantly, impaired cRNAs assembly induces the accumulation of heterochromatic cRNAs and the formation of unscheduled RNA::DNA hybrids (R-loops). Linker histones H1 are known to regulate chromatin structure and compaction and, indeed, we show that dH1 depletion perturbs chromatin organization, reducing nucleosome occupancy and specifically increasing accessibility and 3D interactions within heterochromatin. These perturbations facilitate annealing of cRNAs to the DNA template, enhancing R-loops formation and cRNAs retention at heterochromatin. Altogether, these results unveil the unexpected contribution of linker histones to RNPs assembly and homeostasis of cRNAs.
Project description:Chromosomal RNAs (cRNAs) are a poorly understood fraction of cellular RNAs that co-purify with chromatin. Here we show that, in Drosophila, cRNAs constitute a heterogeneous group of RNA species that cover ~28% of the genome. Intriguingly, we found that cRNAs are highly enriched in heterochromatic transcripts. Our results show that heterochromatic cRNAs interact with the hnRNP A/B proteins hrp36 and hrp48 to assemble into RNP particles. We also show that depletion of linker histone dH1, a major component of chromatin, impairs assembly of hrp36 and hrp48 onto heterochromatic cRNAs. Concomitantly, impaired cRNAs assembly induces the accumulation of heterochromatic cRNAs and the formation of unscheduled RNA::DNA hybrids (R-loops). Linker histones H1 are known to regulate chromatin structure and compaction and, indeed, we show that dH1 depletion perturbs chromatin organization, reducing nucleosome occupancy and specifically increasing accessibility and 3D interactions within heterochromatin. These perturbations facilitate annealing of cRNAs to the DNA template, enhancing R-loops formation and cRNAs retention at heterochromatin. Altogether, these results unveil the unexpected contribution of linker histones to RNPs assembly and homeostasis of cRNAs.