Dimerisation of the PICTS complex via LC8/Cut-up drives co-transcriptional transposon silencing in Drosophila.

Evelyn L Eastwood,Benjamin Czech,Vasileios Frantzis,Kayla A Jara,Gregory J Hannon,Emma Kneuss,Elisar J Barbar,Marzia Munafò,Susanne Bornelöv

doi:10.7554/elife.65557

Evelyn L Eastwood, Benjamin Czech + Show 7 more

Open Access

https://doi.org/10.7554/elife.65557

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Abstract

In animal gonads, the PIWI-interacting RNA (piRNA) pathway guards genome integrity in part through the co-transcriptional gene silencing of transposon insertions. In Drosophila ovaries, piRNA-loaded Piwi detects nascent transposon transcripts and instructs heterochromatin formation through the Panoramix-induced co-transcriptional silencing (PICTS) complex, containing Panoramix, Nxf2 and Nxt1. Here, we report that the highly conserved dynein light chain LC8/Cut-up (Ctp) is an essential component of the PICTS complex. Loss of Ctp results in transposon de-repression and a reduction in repressive chromatin marks specifically at transposon loci. In turn, Ctp can enforce transcriptional silencing when artificially recruited to RNA and DNA reporters. We show that Ctp drives dimerisation of the PICTS complex through its interaction with conserved motifs within Panoramix. Artificial dimerisation of Panoramix bypasses the necessity for its interaction with Ctp, demonstrating that conscription of a protein from a ubiquitous cellular machinery has fulfilled a fundamental requirement for a transposon silencing complex.

Highlights

Large portions of eukaryotic genomes are occupied by repetitive sequences and mobile genetic elements (Cosby et al, 2019)
Ctp associates with the Panoramix-induced co-transcriptional silencing (PICTS) complex and is required for transposon control Previously, we and others showed that Panx, Nuclear export factor 2 (Nxf2), and NTF2-related export protein 1 (Nxt1) form a complex that acts downstream of Piwi target engagement to silence transposons at the transcriptional level (Batki et al, 2019; Fabry et al, 2019; Murano et al, 2019; Zhao et al, 2019)
When considering ChIP-seq signal around all gypsy, mdg1, blood, 297, or 412 insertions in the ovarian somatic cells (OSCs) genome, we detected a significant enrichment for H3K4me2 and depletion of H3K9me3 upon knockdown of PICTS complex components, while control regions were only marginally affected (Figure 2J). This effect is illustrated when looking at a gypsy insertion in the gene expanded on chromosome 2L (Figure 2—figure supplement 1F), which shows that increased transcriptional output across this locus correlates with loss of H3K9me3 and acquisition of H3K4me2 around the gypsy insertion site. These analyses show that Ctp contributes to the repressive chromatin states observed at transposon loci in OSCs, indicating that Ctp is required for the cotranscriptional gene silencing (TGS)

Summary

Introduction

Large portions of eukaryotic genomes are occupied by repetitive sequences and mobile genetic elements (Cosby et al, 2019). Recent work has uncovered a complex consisting of Panoramix (Panx), Nuclear export factor 2 (Nxf2) and NTF2-related export protein 1 (Nxt1) termed PICTS ( known as SFiNX) that functions downstream of Piwi and upstream of the chromatin modifying machinery to effect TGS (Batki et al, 2019; Fabry et al, 2019; Murano et al, 2019; Sienski et al, 2015; Yu et al, 2015; Zhao et al, 2019) While both Panx and Nxf can induce transcriptional repression when artificially recruited to reporters, the link to epigenetic modifiers and the transcriptional silencing activity of the PICTS complex seems to reside within Panx itself (Batki et al, 2019; Fabry et al, 2019). Considered together, our data reveal that Ctp-mediated higher order PICTS complex assembly is essential for heterochromatin formation and silencing

Results

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Discussion

Materials and methods