Mili Interacts with Tudor Domain-Containing Protein 1 in Regulating Spermatogenesis

Abstract

SUMMARY Piwi proteins play essential roles in germline development, stem cell self-renewal, epigenetic regulation, and transposon silencing, and are implicated in oncogenesis [1-7]. Piwi proteins bind to a complex class of small non-coding RNAs called Piwi-Interacting RNAs (piRNAs) [8]. Mammalian Piwi proteins such as Mili are localized in the cytoplasm of spermatogenic cells, where they are associated with a germline-specific organelle called the nuage or its derivative, the chromatoid body, as well as with polysomes [9]. To investigate the molecular mechanisms mediated by Mili, we searched for Mili interacting proteins using co-immunoprecipitation and mass spectrometry. Here we report that Mili specifically interacts with Tudor Domain Containing Protein 1 (Tdrd1; a.k.a. Mouse Tudor Repeat 1, Mtr-1), a germline protein that contains multiple Tudor domains [10, 11]. This RNA-independent interaction is mediated through the N-terminal domain of Mili and the N-terminal region of Tdrd1 containing the Myeloid Nervy DEAF-1 (MYND) domain and first two Tudor domains. In addition, Mili positively regulates Tdrd1 expression at the mRNA level. Furthermore, Mili and Tdrd1 mutants share similar spermatogenic defects. However, Tdrd1, unlike Mili, is not required for piRNA biogenesis. Our results suggest that Mili interacts with Tdrd1 in the nuage and chromatoid body. This interaction does not contribute to piRNA biogenesis, but represents a regulatory mechanism critical for spermatogenesis.

Mouse Piwi interactome identifies binding mechanism of Tdrkh Tudor domain to arginine methylated Miwi

“…Very recently, several groups have characterized the association of specific germline multiTudor proteins with the Piwi family proteins and have confirmed Mili-Tdrd1 and Miwi-Tdrd6 interactions, which coincides with our proteomic data (9,17,18). During the preparation of our manuscript, an elegant study has shown a comprehensive analysis of mouse Piwi complexes and demonstrated that germline Tudor proteins direct critical protein-protein interactions with Piwi proteins that are important for small RNA production and proper operation of the piRNA pathway (30).…”

“…Only one Tudor protein, Tdrd1, was identified as one of the top hits in the Mili complex (Fig. 1B), consistent with recent findings characterizing a Mili-Tdrd1 interaction (17,18). Due to the sensitivity and detection limit of the method, we cannot exclude the possibility that other germline Tudor proteins might also associate with Miwi or Mili complexes.…”

PIWI proteins and PIWI-interacting RNAs function in Hydra somatic stem cells

“…Immuno-electron microscopy demonstrated that both Hywi and Hyli are associated with electron-dense perinuclear structures similar to what is seen in the germlines of several animals, including Drosophila, mice, and zebrafish ( Fig. 1 K and L) (14)(15)(16)(17)(18)(19)(20).…”

Transposable elements in the mammalian germline: a comfortable niche or a deadly trap?

“…This phenotype was originally described for the methylationdeficient Dnmt3L mutant males (Bourc'his and Bestor, 2004), observed again in PIWI mutants (Carmell et al, 2007;Aravin et al, 2007b) and more recently recapitulated in mutations for proteins supporting the piRNA/ PIWI pathway, such as the Tudor domain-containing proteins TDRD1 and TDRD9 (Ollinger et al, 2008;Soper et al, 2008;Ma et al, 2009;Reuter et al, 2009;Shoji et al, 2009). Tudor domain-containing proteins assemble specialized RNA processing platforms in the cytoplasm of germ cells, to which they recruit PIWI proteins through their ability to bind symmetrically dimethylated arginines, a post-translational modification undergone by both MILI and MIWI2 (Aravin et al, 2009;Vagin et al, 2009;Wang et al, 2009;Shoji et al, 2009). As a whole, directed mutagenesis in mice has allowed the identification of nine proteins with a key role in safe guarding the male germline against TEs (Table 1 and Figure 1): Dnmt3L, Dnmt3A, MIWI2, MILI, Maelstrom (MAEL), TDRD1, TDRD9, GASZ and Tex19.1 (Testis expressed 19.1) (Ollinger et al, 2008;Soper et al, 2008;Ma et al, 2009).…”