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Bombyx mori

ABSTRACT: High throughput sequencing of Siwi and BmAGO3 associated piRNAs from Bombyx mori BmN4 cells

PROVIDER: PRJNA251635 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR1334910.fastq.gz	Fastqsanger.gz
	SRR1334909.fastq.gz	Fastqsanger.gz
	SRR1634365.fastq.gz	Fastqsanger.gz
	SRR1634366.fastq.gz	Fastqsanger.gz
	SRR1634367.fastq.gz	Fastqsanger.gz

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High throughput sequencing of Siwi associated piRNAs from Bombyx mori BmN4 cells

Project description:High throughput sequencing of Siwi associated piRNAs from Bombyx mori BmN4 cells

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High throughput sequencing of Siwi and BmAGO3 associated piRNAs from Bombyx mori BmN4 cells

Project description:In germ cells, piRNAs are amplified through the Ping-Pong cycle that depends on reciprocal Slicer-mediated target RNA cleavage by two PIWI members. A germ-specific DEAD-box protein Vasa is required for the process. However, Vasa’s function is poorly understood. Here, we show that target RNAs cleaved by a Bombyx mori (silkworm) PIWI, Siwi, remain to be bound with the protein upon cleavage, but are released in the presence of Vasa in B. mori (BmVasa) and ATP. Under normal conditions, BmVasa co-purifies with Siwi, but not with second B. mori PIWI BmAgo3. However, when BmVasa loses the ATP-binding and RNA-unwinding activities, BmVasa avidly associates with Siwi and BmAgo3, which contains transposon transcripts predominantly in sense orientation, the sources of BmAgo3-piRNAs. Without BmVasa, BmAgo3 is devoid of piRNAs. Thus, BmVasa actively releases target RNAs from Siwi, upon its cleavage, to urge BmAgo3-piRNA complex formation in the Ping-Pong cycle, enabling continuous supply of piRNAs in germ cells.

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Project description:As important roles of small RNA pathways, AGO proteins mediate interaction of incorporated small RNAs with their targets. The resolution of AGO associated small RNAs showed a significant landscape of AGO proteins and their binding small RNAs. To characterize small RNAs that associated with BmAGO2 protein in Bombyx mori, the small RNA population associated with BmAGO2 in BmN cells was extracted from the AGO immunoprecipitated complex and the small RNAs between 17nt to 50nt separated by a polyacrylamide gel electrophoresis were subjected to library construction and deep sequencing.The high throughput sequencing yielded a total of 11691441 reads, representing 813,702 unique reads with a abundance from 5731905 to 1. Small RNA associated with AGO2 of BmN cell infected with ie1-bacmid recombinant viruses were performed by high-throughput using Hiseq 2000.

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