The Xenopus homolog of Bucky

The Xenopus homolog of Bucky ball is known as XVelo (Vegetally localized −1), first identified based on the vegetal pole localization of XVelo RNA (Claussen and Pieler, 2004). A recent study identified XVelo as a Balbiani body enriched protein, and identified prion-like sequences within the amino terminal region of XVelo (Boke et al., 2016) (Fig. 3). In that work, the authors showed that the prion-like domain (PLD) of XVelo was necessary and sufficient for Balbiani body localization of overexpressed XVelo fusion proteins in ovo, and to drive formation of amyloid-like meshworks in in vitro and cell free reconstitution assays (Boke et al., 2016). Notably, with respect to a potential role for the Balbiani body as an oocyte mitochondrial bottleneck discussed above, Boke and colleagues showed that these XVelo meshworks could entrap mitochondria by a mechanism that requires the prion-like domain (Boke et al., 2016). Although the amino Phenyl sulfate level conservation between XVelo and zebrafish Bucky ball is poor over much of the sequence, the proteins are most similar in their amino terminal region (Bontems et al., 2009; Boke et al., 2016), which includes the PLD. Evidence for a conserved activity includes the ability of both proteins to drive assembly of structures that can recruit mitochondria and RNAs (Marlow and Mullins, 2008; Bontems et al., 2009; Heim et al., 2014), and experiments in which versions of XVelo and Bucky ball with swapped PLD domains supported stable matrix formation in photobleaching and recovery assays (Boke et al., 2016). Interestingly, versions of XVelo containing the PLD, but lacking the C-terminal region could still form a matrix in overexpression assays, but could not recruit mitochondria to the matrix (Boke et al., 2016). Therefore, the PLD is sufficient for matrix formation, but the C-term is required to coordinate mitochondrial recruitment either directly or indirectly. Notably, both of the previously characterized zebrafish bucky ball mutant alleles that encode for truncated Bucky ball proteins (if any protein is produced) lacking the entire C-terminus or only the last few dozen amino acids of the protein fail to support Balbiani body formation and recruitment of mitochondria and RNA (Marlow and Mullins, 2008; Bontems et al., 2009). Moreover, consistent with an important role for the C-terminus in recruiting mitochondria and Balbiani body proteins, the overexpression phenotypes of transgenic animals expressing Bucky ball, and rescue of the mitochondria localization and Balbiani body formation defects of bucky ball mutants require the Bucky ball C-terminus in transgenic (Heim et al., 2014) and transient rescue assays (Bontems et al., 2009). Interestingly, Boke and colleagues identified residues in the C-terminus of XVelo that lack a known RNA recognition consensus sequence, but have characteristics that might confer RNA binding properties similar to reports for Drosophila Oskar, another intrinsically disordered protein with essential roles in germ cell formation (Jeske et al., 2015). Consistent with XVelo meshworks supporting RNA recruitment, which is a previously described function of zebrafish Bucky ball (Marlow and Mullins, 2008; Bontems et al., 2009; Heim et al., 2014), versions of XVelo with the prion-like domain but lacking the rest of the protein formed meshworks, but did not recruit a labeled nanos/xcat-2 RNA there (Boke et al., 2016). Both XVelo and Bucky ball, like Oskar, have previously been shown to bind RNA binding proteins (Heim et al., 2014; Boke et al., 2016; Jeske et al., 2015). Thus, it appears that a potential shared feature of the germ plasm assemblers may be the capacity to recruit RNAs by two mechanisms – direct interaction via their putative RNA binding domains and through their interactions with RNA binding proteins. However, in the case of XVelo it remains to be determined whether recruitment of nanos/xcat-2 to the network occurs via direct or indirect RNA interactions, as both XVelo and Bucky ball have been shown to interact with the conserved RNA binding protein, Rbpms2 (RNA binding protein with multiple splice isoforms), which binds to Balbiani body localized germ plasm RNAs such as bucky ball and nanos/xcat-2 (Heim et al., 2014; Nijjar and Woodland, 2013; Song et al., 2007; Aguero et al., 2016). Notably, XVelo/Bucky ball interactions with Rbpms2 protein are mediated via the XVelo/Bucky ball N-terminus (Heim et al., 2014; Nijjar and Woodland, 2013), which is also the region harboring the prion-like domain (Boke et al., 2016); however, whether interaction with Rbpms2 promotes or represses Buc/XVelo functions in Balbiani body and matrix formation, is involved in bucky ball RNA regulation, or only facilitates recruitment of RNAs to the assembled structure remains to be determined.