The conserved zinc-finger protein GTSF1 helps PIWI proteins achieve their full catalytic potential

Abstract

Argonaute proteins use nucleic acid guides to find and bind specific DNA or RNA target sequences. Argonaute proteins can be found in all kingdoms of life, and play diverse biological functions including genome defense, gene regulation, and chromosome partitioning. Many Argonautes retain their ancestral endoribonuclease activity, cleaving the phosphodiester bond between target nucleotides t10 and t11. In animals, a specialized class of Argonautes, the PIWI proteins, use 21–35 nt PIWI-interacting RNAs (piRNAs) to direct transposon silencing, protect the germline genome, and regulate gene expression during gametogenesis1. The piRNA pathway is required for fertility in one or both sexes of nearly all animals. Both piRNA production and function require RNA cleavage catalyzed by PIWI proteins. Spermatogenesis in mice and other placental mammals requires three distinct, developmentally regulated PIWI proteins: MIWI (PIWIL1), MILI (PIWIL2), and MIWI2 (PIWIL4)2 4; the piRNA-guided endoribonuclease activities of MIWI and MILI are essential to produce functional sperm5,6. piRNA-directed silencing in mice, insects, and worms also requires Gametocyte-Specific Factor 1 (GTSF1), a PIWI-associated protein of unknown function7 12. Here, we report that GTSF1 potentiates the weak, intrinsic, piRNA-directed RNA cleavage activities of PIWI proteins, transforming them into efficient endoribonucleases. GTSF1 represents the first example of an auxiliary protein that potentiates the catalytic activity of an Argonaute protein.

Developmental roles and molecular mechanisms of Asterix/GTSF1

“…This connection—from Asterix to tRNAs to LTR transposons—is reinforced by genetic observations that LTR retrotransposons are disproportionately affected by Asterix/GTSF1 depletion (Donertas et al, 2013; Ipsaro et al, 2021; Muerdter et al, 2013; Ohtani et al, 2013). Moreover, the same preprint that indicates mouse GTSF1 potentiation of piRISC catalysis also reported an increase in enzymatic activity for sponge GTSF1 with sponge Piwi (EfPIWI) in a tRNA‐containing buffer (Arif et al, 2021). Together, these associations suggest that GTSF1 could be enhancing piRNA‐directed silencing is through tRNA‐directed transposon recognition.…”

“…Mature piRISCs engage with target transcripts (top) and enforce silencing by endonucleolytic “slicing”. This activity is native to mammalian Piwi proteins like MILI, but is potentiated by GTSF1 (Arif et al, 2021). Recent data suggest that recognition of target loci by GTSF1 is accomplished through tRNA‐meditated interactions with GTSF1 and the target transcript (Ipsaro et al, 2021).…”

“…However, those complexes do not seem to participate in ping‐pong to the same extent. Moreover, interactions between sets of Piwi and GTSF1 paralogs do appear to potentiate cleavage universally, although this is the case for GTSF1 with MIWI and MILI (Arif et al, 2021)…”

“…Similar assertions have been made in mammalian systems, where it has been posited that mouse GTSF1 is required for MIWI2‐piRISCs to stabilize and/or grasp target RNAs (Yoshimura et al, 2018). Additionally, a recent preprint suggests that mouse GTSF1 is a factor that potentiates mouse Piwi proteins' catalysis, increasing target cleavage by orders of magnitude (Arif et al, 2021).…”

“…In addition to its interactions with Piwi proteins, RNA interactions are also critical to Asterix/GTSF1 function (Arif et al, 2021; Ipsaro et al, 2021). Studies of recombinant mouse GTSF1 were the first to show robust and specific co‐precipitation GTSF1 with RNA, which were specifically identified as tRNAs.…”