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

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.

Rewiring of chromatin state and gene expression by transposable elements

“…(b) Hypothetical model showing the condensation of multiple Piwi–piRNA silencing complexes. Possible formation of LLPS is suggested in recent studies (Eastwood et al., 2021; Schnabl et al., 2021)…”

“…Nxf2 associates with the nascent transcript of target TEs to form a silencing complex. Recent studies have suggested that Panx can further associate with Cut‐up/LC8 and form a dimerized structure, suggesting the possibility that the Piwi–piRNA silencing complex can form a liquid‐liquid phase separation (LLPS) (Eastwood et al., 2021; Schnabl et al., 2021) (Figure 4b). HP1a, which is recruited to H3K9me, and the linker histone H1 are further associated with the piRNA target regions to induce a stable heterochromatin structure (Huang et al., 2013; Iwasaki et al., 2016; Le Thomas et al., 2013; Rozhkov et al., 2013).…”

“…Similarly, in the case of Piwi–piRNA‐mediated silencing, chromatin regulation upon heterochromatin formation has been shown to spread for several tens of kilobases from target TEs (Sienski et al., 2012). Recent studies have suggested the possible formation of an LLPS upon Piwi–piRNA‐mediated silencing of TEs (Eastwood et al., 2021; Schnabl et al., 2021). Therefore, piRNA target regions and their silencing machinery may form a spatial complex to silence their targets.…”

Maternally inherited piRNAs direct transient heterochromatin formation at active transposons during early Drosophila embryogenesis

“…Piwi in embryos showed high levels of antisense piRNAs targeting roo (~16% or all TE-targeting reads) and 297 (~9%), consistent with an ability of maternally deposited Piwi to potentially recognize these TEs when expressed in the embryo. In Drosophila ovaries, coTGS depends on Piwi-mediated recruitment of the PICTS/SFiNX complex and correlates with the deposition of H3K9me3 marks at TE insertions and surrounding genomic regions (Batki et al, 2019;Eastwood et al, 2021;Fabry et al, 2019;Murano et al, 2019;Schnabl et al, 2021;Sienski et al, 2015;Yu et al, 2015;Zhao et al, 2019). Due to the poor conservation of the genomic locations of transposon insertions between different Drosophila strains, we used whole genome sequencing to de novo identify the TE insertion sites present in our control w 1118 flies (see methods).…”

Dynein light chain-dependent dimerization of Egalitarian is essential for maintaining oocyte fate in Drosophila

“…Recently, Panoramix (Panx), a protein that functions in the piRNA pathway was also shown to dimerize in a Dlc-dependent manner (Eastwood et al, 2021;Schnabl et al, 2021). Artificial dimerization of a panx mutant that could no longer interact with Dlc (using the same GCN4 leucine zipper) restored the function of panx (Eastwood et al, 2021;Schnabl et al, 2021).…”