The piRNA pathway maintains genome integrity by silencing transposons cotranscriptionally in the nucleus through recognition of nascent transposon RNAs and recruitment of endogenous transcriptional and chromatin-level repressive mechanisms to transposon loci. However, the molecular link between the SFiNX complex, which recognizes nascent transposon RNA, and downstream effector complexes has remained elusive. Here, we demonstrate that the Small ovary (Sov) protein mediates this connection. By mapping the functional activities of its structural elements, we reveal that Sov contributes to transposon silencing through two distinct molecular mechanisms. First, Sov specifically directs nascent transposon transcripts toward nuclear RNA exosome-mediated degradation by physically interacting with the RNA decay factor TEsup1. Second, Sov directly binds the heterochromatin protein HP1a via multiple conserved motifs and undergoes phase separation, facilitating heterochromatin formation and genome-wide gene repression. Genetic analyzes of sov mutants reveal that these functions are separable: RNA-mediated transcriptional silencing is essential for piRNA pathway activity, while phase separation-dependent heterochromatin regulation is critical for stable transposon repression. We propose that Sov acts as a molecular scaffold in piRNA-guided transposon silencing, integrating transposon recognition with cotranscriptional RNA decay and chromatin-based regulatory pathways.
Z. Földi, Melinda Bence, Zsanett Takács et al.· bioRxiv· 0 citations
Assemblysomes are ribosome-nascent chain condensates that regulate co-translational processes through liquid–liquid phase separation, yet the sequence determinants underlying their formation remain incompletely understood. Previous studies identified the DNA helicase Sgs1 as an assemblysome-associated protein; however, whether its N-terminal region possesses intrinsic phase separation propensity has not been experimentally examined. Here, we investigated the first 135 amino acids of Sgs1 using a light-inducible optoDroplet assay. A mCherry–Cry2–Sgs11–135 fusion construct was compared with the established positive control FUS–mCherry–Cry2 and the negative control mCherry–Cry2 in live HEK293T cells. Following blue-light activation, Sgs11–135 reproducibly formed reversible condensates, indicating intrinsic phase separation propensity. Quantitative image analysis revealed light-dependent increases in condensate number, average condensate area, and integrated condensate fluorescence intensity. Compared with FUS, Sgs11–135 formed slightly fewer and smaller condensates but displayed reproducible light-dependent condensate formation. These findings indicate that the Sgs1 N-terminal region exhibits intrinsic phase separation propensity in a validated optogenetic assay. Although this proof-of-principle study does not establish the molecular mechanism of assemblysome formation, the results are consistent with the hypothesis that the Sgs1 N-terminus may contribute to the multivalent interactions underlying assemblysome organization.
Bence György Gombás, Erika Gábor, V. Honti et al.· Biomolecules· 0 citations
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