Rapid repetitive element-mediated expansion of piRNA clusters in mammalian evolution

Abstract

Piwi-interacting RNAs (piRNAs) are approximately 30 nucleotide noncoding RNAs that may be involved in transposon silencing in mammalian germline cells. Most piRNA sequences are found in a small number of genomic regions referred to as clusters, which range from 1 to hundreds of kilobases. We studied the evolution of 140 rodent piRNA clusters, 103 of which do not overlap protein-coding genes. Phylogenetic analysis revealed that 14 clusters were acquired after rat-mouse divergence and another 44 after rodent-primate divergence. Most clusters originated in a process analogous to the duplication of protein-coding genes by ectopic recombination, via insertions of long sequences that were mediated by flanking chromosome-specific repetitive elements (REs). Source sequences for such insertions are often located on the same chromosomes and also harbor clusters. The rate of piRNA cluster expansion is higher than that of any known gene family and, in contrast to other large gene families, there was not a single cluster loss. These observations suggest that piRNA cluster expansion is driven by positive selection, perhaps caused by the need to silence the ever-expanding repertoire of mammalian transposons.

Fig. 1.

Acquisition of a cluster-harboring sequence. Alignment of a cluster-harboring segment (yellow) in sister 1 to a gap in sister 2 and in an outgroup indicates that this segment was acquired in the lineage of sister 1 after it diverged from the lineage of sister 2. Flanking protein-coding genes are depicted in orange and green.

Fig. 2.

Schematic used to identify ectopic recombination as the mechanism of an insertion. (A) Architecture of a typical cluster-harboring genomic region. Two protein-coding genes (orange and green) flank an intergenic region containing an acquired cluster (blue) that is preceded by a RE (red). The inserted segment is depicted within brackets. (B) The inserted segment (Left) is scanned against the genome to locate the source paralog, which is depicted within brackets in its genomic context. The source paralog harbors a paralogous cluster (blue) and is preceded by a RE (Right). (C) Alignment of the cluster- and source paralog-harboring regions indicates that similarity between the 2 sequences includes the REs preceding paralogous clusters. (D) The most likely insertion mechanism is ectopic recombination. Following a double-stranded break (Bottom strand), recombination occurs between homologous REs preceding the source paralog (Top strand) and the site of the double-stranded break. Extension and reannealing of the broken strand generates a new cluster-harboring segment.