The multiple Tudor domain-containing protein TDRD1 is a molecular scaffold for mouse Piwi proteins and piRNA biogenesis factors

Abstract

Piwi-interacting RNAs (piRNAs) are small noncoding RNAs expressed in the germline of animals. They associate with Argonaute proteins of the Piwi subfamily, forming ribonucleoprotein complexes that are involved in maintaining genome integrity. The N-terminal region of some Piwi proteins contains symmetrically dimethylated arginines. This modification is thought to enable recruitment of Tudor domain-containing proteins (TDRDs), which might serve as platforms mediating interactions between various proteins in the piRNA pathway. We measured the binding affinity of the four individual extended Tudor domains (TDs) of murine TDRD1 protein for three different methylarginine-containing peptides from murine Piwi protein MILI. The results show a preference of TD2 and TD3 for consecutive MILI peptides, whereas TD4 and TD1 have, respectively, lower and very weak affinity for any peptide. The affinity of TD1 for methylarginine peptides can be restored by a single-point mutation back to the consensus aromatic cage sequence. These observations were confirmed by pull-down experiments with endogenous Piwi and Piwi-associated proteins. The crystal structure of TD3 bound to a methylated MILI peptide shows an unexpected orientation of the bound peptide, with additional contacts of nonmethylated residues being made outside of the aromatic cage, consistent with solution NMR titration experiments. Finally, the molecular envelope of the four tandem Tudor domains of TDRD1, derived from small angle scattering data, reveals a flexible, elongated shape for the protein. Overall, the results show that TDRD1 can accommodate different peptides from different proteins, and can therefore act as a scaffold protein for complex assembly in the piRNA pathway.

Tdrd12 Is Essential for Germ Cell Development and Maintenance in Zebrafish

“…Loss of Tdrd1 leads to defective nuage structures in germ cells, transposon desilencing, and the loss of germ cells in zebrafish. These observations also demonstrated the similar functions shared between zebrafish Tdrd1 and its mouse counterpart [21,26,27]. Tdrd6 is required for normal primordial germ cell formation and the accumulation of maternally inherited piRNAs in zebrafish.…”

A Structural Perspective on Readout of Epigenetic Histone and DNA Methylation Marks

“…These studies have shed light on the principles underlying recognition of the symmetrical dimethylated arginine mark (Rme2s) (Liu et al 2010a;Liu et al 2010b;Mathioudakis et al 2012). The structures of complexes containing other Tudor group proteins led to similar conclusions about the nature of expanded Tudor domain interactions with methylated arginines (Liu et al 2010a;Liu et al 2010b;Mathioudakis et al 2012). The results of the complex between the SND1 (staphylococcal nuclease domain-containing 1) extended Tudor module and the R14me2s-containing amino-terminal PIWI peptide is discussed here (Liu et al 2010b).…”

The piRNA Pathway Guards the Germline Genome Against Transposable Elements

“…Tudor family proteins containing the eponymous Tudor domain can bind methylated arginines in PIWI proteins, and this relationship appears to be conserved in many species. Many Tudor proteins have several Tudor domains and act as a scaffold for the formation of higher-order complexes (Huang et al 2011a; Mathioudakis et al 2012). In germ cells of Drosophila and in mouse testis, this mode of protein interaction translates into formation of distinct perinuclear granules similar to P-bodies, which harbour components of the piRNA pathway and have been known for a long time in cell biology as “nuage” (Aravin et al 2008, 2009; Brennecke et al 2007; Malone et al 2009).…”