HIWI cDNA was first partially isolated as an ortholog of Drosophila PIWI (Cox et al., 1998) and then fully identified in a human testis cDNA library (Qiao et al., 2002; Sharma et al., 2001).

HIWI has two transcript variants. Variant 1 (NCBI Ref Seq: NM_004764.4) was cloned from a human testis cDNA library (Qiao et al., 2002; Sharma et al., 2001). It is 3591 nt in length and comprises 21 verified exons, of which 20 are coding exons with a total length of 2586 nt. The 5- and 3-UTRs are 271 and 734 nt long, respectively. Variant 2 has an alternative 3-UTR, resulting in shorter protein with truncated C-terminus compared with variant 1. Variant 2 was detected by transcriptome analyses of the "full-length long JAPAN" (FLJ) collection (Ota et al., 2004).

HIWI protein (NCBI Ref Seq: NP_004755.2) belongs to the PIWI subgroup of Argonaute family proteins, evolutionarily conserved proteins containing two characteristic protein motifs, PAZ and PIWI (Carmell et al., 2002). Argonaute family proteins can be divided into two subclades, AGO and PIWI, and form complexes with small RNAs to regulate gene expressions (Carmell et al., 2002; Hock and Meister, 2008).

The HIWI gene encodes an 861-amino-acid protein (98 kDa). The HIWI protein contains two characteristic protein motifs: a PAZ domain (aa 278-413) and a PIWI domain (aa 556-847). In Argonaute family proteins, the PAZ domain serves as a docking site for the 3-end of small RNA, whereas the PIWI domain has a structure similar to RNaseH, which cleaves the RNA strand of an RNA-DNA hybrid (Elkayam et al., 2012; Parker and Barford, 2006). Indeed, all three species of the Drosophila PIWI protein family show small RNA-guided RNA cleavage (slicer) activity (Gunawardane et al., 2007; Nishida et al., 2007; Saito et al., 2006). In mouse, MIWI (mouse orthologs of HIWI) and MILI also have the slicer activity, which is essential for transposon silencing and fertility (De Fazio et al., 2011; Reuter et al., 2011). In addition, the HIWI protein contains glycine-arginine-rich repeats mainly in its N-terminus (aa 3-15:GRARARARGRARG; aa18-20:GRG; aa728-730:GRG), and these are the motifs for symmetrical dimethylarginine (sDMA) modifications catalyzed by PRMT5 methyltransferase (Kirino et al., 2009). sDMA positions of MIWI were determined by mass spectrometry (Chen et al., 2009; Vagin et al., 2009). PIWI sDMAs serve as binding elements for TUDOR-domain containing proteins, and the sDMA-dependent protein interactions play crucial roles in piRNA function and germline development (Chen et al., 2011; Siomi et al., 2010).

Expression of the PIWI protein family is restricted to germline cells (Farazi et al., 2008).

PIWI protein family was named after the Drosophila protein PIWI (P-element induced wimpy testis) (Carmell et al., 2002; Farazi et al, 2008). The Drosophila PIWI was first identified in a genetic screen for mutants that affect stem cell division in the germline, and subsequent studies demonstrated that Drosophila PIWI is essential for gametogenesis and is a key regulator of female germline stem cells (Cox et al., 1998; Cox et al., 2000; Lin and Spradling, 1997). PIWI protein mutations in various organisms, including mice and zebrafish, commonly cause defects in gametogenesis, indicating evolutionarily conserved essential roles for PIWI proteins in germline development (Carmell et al., 2007; Deng and Lin, 2002; Houwing et al., 2008; Houwing et al., 2007; Kuramochi-Miyagawa et al., 2004). Since 2006, the small RNAs bound to PIWI proteins have been identified and termed PIWI-interacting RNAs (piRNAs) (Siomi et al., 2011). piRNAs are 24-31 nt in length and are a highly complex mix of sequences derived from defined genomic regions called piRNA clusters. Many piRNAs are derived from transposable elements, and PIWI/piRNA complexes play a crucial role in silencing transposons during germline development (Siomi et al., 2011).