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Global coupling of R-loops with transcription dynamics modulates early development and survival of Drosophila

ABSTRACT: Global coupling of R-loops with transcription dynamics modulates early development and survival of Drosophila

PROVIDER: PRJNA1124981 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR29442759_1.fastq.gz	Fastqsanger.gz
	SRR29442759_2.fastq.gz	Fastqsanger.gz
	SRR29442760_1.fastq.gz	Fastqsanger.gz
	SRR29442760_2.fastq.gz	Fastqsanger.gz
	SRR29442761_1.fastq.gz	Fastqsanger.gz

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Global coupling of R-loops with transcription dynamics modulates early development and survival of Drosophila

Project description:R-loops are involved in many biological processes in cells, yet the regulatory principles for R-loops in vivo and their impact on development remain to be explored. Here, we modified the CUT&Tag strategy to profile R-loops in Drosophila at multiple developmental stages. While high GC content promotes R-loop formation in mammalian cells, it is not required in Drosophila. In contrast, RNAPII abundance on chromatin appears to be a universal inducing factor for R-loop formation in the Drosophila genome, including active promoters and enhancers, and H3K27me3 decorated repressive regions and intergenic repeat sequences. Importantly, such a regulatory relationship is dynamically maintained throughout development and many development-related transcription factors that may regulate RNAPII activation and R-loop dynamics were uncovered. By ablating the elongation factor Spt6, we further showed the global R-loop induction coupled with RNAPII pausing, genome instability, and defects in larvae survival and cuticle development. Overexpression of rnh1 largely reversed the upregulation or downregulation of gene expression depending on gene length, DNA damage, and defects in survival and cuticle development. Altogether, our findings indicate that dynamic R-loop regulation is dictated by RNAP pausing and transcription activity, and plays a feedback role in gene regulation, genome stability maintenance, and Drosophila development.

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Project description:The R-loop in the genome participates in modulating transcription, replication and DNA damage repair, yet its regulation and biologic impact on development in vivo remain to be explored. Here, we modified CUT&Tag strategy and established R-loop profiles of Drosophila across multiple developmental stages. While lack of GC preference, the broad class of R-loops in Drosophila all highly follow the RNAPII pausing, which is also a conserved R-loop regulatory mechanism in mammals. Meanwhile, RNAPII-regulated R-loop may be regulated by multiple developmental specificity and universality transcription factors. To test the specificity and universality of transcription factor-mediated R-loop involvement in development, we knocked out the transcription elongation factor Spt6, upregulated R-loop and increased RNAPII pausing, and altered the expression of critical genes and cutin genes, thereby affecting Drosophila lifespan and cutin development. Interestingly, complementation of rnh1 could partially rescue the transcription factor deletion phenotype. We conclude that RNAPII is a conserved mechanism of R-loop regulation across species and organism specific damage caused by R-loop dysregulation is mediated by the functions of transcription factors.

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