Tejas functions as a core component of nuage assembly and precursor processing inDrosophilapiRNA biogenesis

Abstract

Piwi-interacting RNAs (piRNAs), a class of 23- to 29-nt gonad-specific small RNAs, function to combat transposons in gonads. piRNAs are thought to be processed and amplified in membrane-less granules called nuage in germline cells. In Drosophila, two PIWI family proteins, several Tudor-domain containing (Tdrd) proteins and RNA helicases are assembled at perinuclear region of germline cells, forming nuage to process into piRNAs. Among those, Tejas (Tej), a fly homolog of mouse Tdrd5, has been known as a robust nuage component required for piRNA biogenesis in germline cells, yet its molecular functions remained elusive. To understand its molecular basis on nuage assembly and functions for piRNA biogenesis, we investigated subcellular localization of fluorescent-tagged nuage proteins including Tej and monitored the behavior of piRNA precursors. Tej functions as a core component for assembly of Vasa and Spindle-E to nuage granules through distinct motifs, respectively. The loss of Tej function resulted in malformation of nuage and accumulation of piRNA precursors en route in processing, perturbing further piRNA biogenesis in germline cells. Our study also revealed that the low complexity region of Tej regulates the mobility of nuage by phase separation. Collectively, we propose that Tej plays a pivotal role in processing of piRNA precursors by assembling RNA helicases, Vasa and Spindle-E, to nuage, by controlling the dynamics of nuage components.

Molecular insight into theDrosophilapiRNA pathway network through a combination of systematic protein interaction screening and structural prediction

“…Instead, we discovered that the Tejas – Krimper complex may provide a direct link between Vasa and Aub. Together with our and recent data (Lin et al 2023) on the Tejas-Spindle E interaction, we conclude that Tejas is a critical core component of nuage.…”

“…Tejas has also been suggested to associate with the TDRDs Tapas ( Drosophila ortholog of TDRD7) and Spindle E (a.k.a. Homeless; Drosophila ortholog of TDRD9) (Patil and Kai 2010; Patil et al 2014; Lin et al 2023), and we confirmed Tapas ( Figure 1B, Supplementary Figure 2P ) and Spindle E as Tejas binding partners ( Figures 1B and 5B, Supplementary Figure 2 ). Because we did not obtain a high confidence structural model for the Tejas - Tapas complex, we did not further characterize this interaction.…”

“…The Tejas mutation V347E had no effect on the Tejas - Spindle E interaction in ReLo assays ( Figure 5I ), suggesting that the SIR can be refined to aa 315-335 of Tejas. An identical structural model of the Tejas-Spindle E complex was also recently reported using AlphaFold2 (Lin et al 2023). In addition to Tejas, Spindle E has been proposed to interact with the TDRDs Tapas (Patil et al 2014) and Qin (TDRD4) (Anand and Kai 2012) and with the RNA helicase Vasa (Pek and Kai 2011), of which we could only confirm Qin as an additional Spindle E interaction partner ( Figures 1B and 5J, Supplementary Figure 2 ).…”

“…The Tejas mutation V347E had no effect on the Tejas -Spindle E interaction in ReLo assays (Figure 5I), suggesting that the SIR can be refined to aa 315-335 of Tejas. An identical structural model of the Tejas-Spindle E complex was also recently reported using AlphaFold2 (Lin et al 2023).…”