Rhino defines H3K9me3-marked piRNA clusters

piRNAs (PIWI-interacting RNAs), which are generally 24–32 nucleotides in length, are defined by their specific association to the PIWI subfamily of AGO/ PIWI family proteins. piRNAs are mainly made in animal gonads, and are essential to silence transposons during germline development [1]. In Drosophila, the genomic regions that transcribe piRNA precursors are named as piRNA clusters. The primary piRNAs are transcribed from piRNA clusters. Most of the piRNA clusters are located in pericentromeric and subtelomeric heterochromatin regions coated with H3K9me3 mark. There are two types of piRNA clusters in the Drosophila ovaries: uni-strand and dual-strand, which are mainly active in somatic follicle cells and germ cells, respectively. The mechanisms defining the uni-strand or dual-strand piRNA clusters are different. Recent studies have shown that Rhino, a germline-specific protein required for primary piRNA production in germline cells, is anchored to H3K9me3-marked chromatin and specifically binds to dual-strand clusters [2, 3]. Rhino belongs to the heterochromatin protein 1 (HP1) family, also called HP1D. Like other proteins in the HP1 family, Rhino contains two conserved domains: N-terminal chromodomain (CD) and C-terminal chromoshadow domain (CSD), and a hinged region in between. The chromodomain is in charge of recognizing the H3K9me3 marker, and the chromoshadow domain is involved in interaction with other proteins. The hinged region in Rhino is much longer than in other members of this family. Here, the molecular features that distinguish Rhino from other HP1 proteins in Drosophila and allow it to specifically recognize dual-strand piRNA clusters are found. We and Le Thomas et al. have recently reported the high resolution structure of Rhino chromodomain (Rhino-CD) in complex with H3K9me3 [4, 5]. Surprisingly, the Rhino-CD forms a dimer both in crystal and in solution, which is unique among all known chromodomains. Each monomer has an aromatic cage binding H3K9me3, a common feature shared by the chromodomains from other HP1 family proteins. But the dimeric Rhino-CD allows the binding of two H3K9me3 peptides in anti-parallel manner. Both aromatic cages and dimerization of Rhino-CD were found to be important to its function both in vitro and in vivo. The Rhino-CD mutants lost binding of H3Kc9me3-labeled polynucleosomes, a mimic of the H3K9me3-polynucleosomes. The corresponding mutation leads to transposon activation and fly sterility [4]. Considering that Rhino specifically locates to dual-strand clusters rather than uni-strand clusters, a special chromatin structures is suggested. Yamanaka et al. have recently proposed a possible link between chromatin boundaries and piRNA clusters …