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SRX5709937: GSM3730468: RNAseq_dNxf2_het_Panx_FL10B_rep1; Drosophila melanogaster; RNA-Seq
1 ILLUMINA (HiSeq X Ten) run: 12.9M spots, 1.9G bases, 720.8Mb downloads

Submitted by: NCBI (GEO)
Study: A Pandas complex adapted for piRNA-guided transposon silencing (RNA-seq)
show Abstracthide Abstract
The repression of transposons by the Piwi-interacting RNA (piRNA) pathway is essential to protect animal germ cells. In Drosophila ovaries, Panoramix (Panx) enforces transcriptional silencing by binding to the target-engaged Piwi-piRNA complex, although the precise mechanisms by which this occur remain elusive. Here, we show that Panx functions together with a germline specific paralogue of a nuclear export factor, dNxf2, and its cofactor dNxt1 (p15) as a ternary complex to suppress transposon expression. Structural and functional analysis demonstrate that dNxf2 binds Panx via its UBA domain and play an important role in transposon silencing through binding to transposon transcripts directly. Unexpectedly, dNxf2 interacts directly with dNxf1 (TAP), which is also essential for transposon silencing. Transient tethering of dNxf2 to nascent transcripts leads to their nuclear retention. Therefore, we propose that dNxf2 may function as a Pandas (Panoramix dNxf2 dependent TAP/p15 silencing) complex, which counteracts the canonical RNA exporting machinery (TAP/p15) and restricts transposons to nuclear peripheries. Our finding may have broader implications for understanding how RNA metabolism modulates epigenetic gene silencing and heterochromatin formation. Overall design: Examination of transcriptome from dNxf2 mutant comparing to dNxf2 heterogygous with panx tethered to FL10B tethered, and Panx mutant comparing to Panx heterogygous with dNxf2 tethered to FL10B from Drosophila ovaries.
Sample: RNAseq_dNxf2_het_Panx_FL10B_rep1
SAMN11463383 • SRS4649108 • All experiments • All runs
Library:
Instrument: HiSeq X Ten
Strategy: RNA-Seq
Source: TRANSCRIPTOMIC
Selection: cDNA
Layout: SINGLE
Construction protocol: Total RNA was prepared with Trizol reagent (Invitrogen), Transcriptome libraries were prepared according (Armour et al. 2009, Nat Methods) using not-so-random priming (NSR). 1 ug total RNA was revere transcribed using SuperScript III enzyme with first-strand NSR primer. RNA template was removed with RNase H (Invitrogen). Then cDNA was mixed with exo-Klenow fragment (NEB) second-strand NSR primer to synthesize the second strand. For PCR amplification, purified second-strand synthesis reaction were mixed with LA Taq Version 2.0 (Takara) and P5-SBS3T-NSR and P7-SBS8N-NSR primers. Products were run on a 2% low-melt agarose gel, and the 250–500 bp range were purified. RNA-seq was performed as described (Vagin et al., 2013 RNA and Armour et al., 2009 Nat Methods). Samples were sequenced on Illumina platforms with HiSeq X Ten.
Experiment attributes:
GEO Accession: GSM3730468
Links:
Runs: 1 run, 12.9M spots, 1.9G bases, 720.8Mb
Run# of Spots# of BasesSizePublished
SRR892899212,924,2961.9G720.8Mb2019-08-14

ID:
7691881

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