Two Piwi proteins, Xiwi and Xili, are expressed in the Xenopus female germline

Abstract

The Argonaute superfamily is a large family of RNA-binding proteins involved in gene regulation mediated by small noncoding RNA and characterized by the presence of PAZ and PIWI domains. The family consists of two branches, the Ago and the Piwi clade. Piwi proteins bind to 21-30-nucleotide-long Piwi-interacting RNAs (piRNAs), which map primarily to transposons and repeated sequence elements. Piwi/piRNAs are important regulators of gametogenesis and have been proposed to play roles in transposon silencing, DNA methylation, transcriptional silencing, and/or post-transcriptional control of translation and RNA stability. Most reports to date have concentrated on the Piwi family members in the male germline. We have identified four Piwi proteins in Xenopus and demonstrate that two, namely, Xiwi1b and Xili, are expressed in the oocyte and early embryo. Xiwi1 and Xili are predominantly found in small, separate complexes, and we do not detect significant interaction of Piwi proteins with the cap-binding complex. Putative nuclear localization and export signals were identified in Xiwi1 and Xili, supporting our observation that Xiwi1, but not Xili, is a nucleo-cytoplasmic protein. Furthermore, by immunoprecipitation of small RNAs, we establish Xiwi1 as a bona fide Piwi protein. These results suggest that the Piwi/piRNA pathway is active in translationally repressed oocytes. This is a significant finding as the Xenopus model provides an excellent tool to study post-transcriptional mechanisms.

Identification of germ plasm‐associated transcripts by microarray analysis of Xenopus vegetal cortex RNA

“…Very little is known about transposon silencing in Xenopus . The Piwi proteins, which have been implicated in multiple aspects of germ cell specification and transposon silencing (Houwing et al, 2007), are expressed in Xenopus eggs (Wilczynska et al, 2009) and a Xenopus Piwi homolog (Xiwi) is enriched in the germ plasm (Lau et al, 2009). Also, the small RNAs bound by Piwi proteins (piRNAs) represent the major class of small RNA in Xenopus oocytes (Lau et al, 2009).…”

Abundant and dynamically expressed miRNAs, piRNAs, and other small RNAs in the vertebrate Xenopus tropicalis

“…As shown in Figure 1B, we detected expression of all four genes thought to be involved in miRNA biogenesis and function (dicer1, rnasen, eif2c2, eif2c4) in all tissues tested. We recently demonstrated that at least two Argonaute proteins of the Piwi family, piwil1.1/piwil1.2 (also known as Xiwi1a/Xiwi1b), and piwil2 (also known as Xili), are present in Xenopus oocytes (Wilczynska et al 2009). We detected piwil1.2 and piwil2 mRNA in both the germline and the soma; however, no Piwi proteins were detected in the soma (Supplemental Fig.…”

“…1A). Third, we previously demonstrated that one Piwi protein of X. tropicalis, piwil1.1/piwil1.2, associated specifically with ;30 nt RNAs in Xenopus oocytes (Wilczynska et al 2009). Next, we validated two of these candidate piRNAs, named piR-1 and piR-2, using Northern blotting.…”

XenopusNanos1 is required to prevent endoderm gene expression and apoptosis in primordial germ cells

“…Neither a mutant of nanos1 with 5Ј sequences non-complementary to the MO (nanos1-mut) nor a control MO with five base changes (Nanos1-Ctrl-MO) had any effect on nanos1 translation (data not shown). We have previously shown that during early developmental stages, Nanos1 is found within germ plasm, colocalizing with the germ plasm marker Xiwi (Lai et al, 2011;Lau et al, 2009;Wilczynska et al, 2009). Importantly, whereas Xiwi was readily detected, Nanos1 protein was not observed in embryos previously injected with Nanos1-MO (20 ng) after staining with anti-Nanos1 and anti-Xiwi antibodies (Fig.…”