Comparative analysis of piRNA sequences, targets and functions in Caenorhabditis nematodes

Piwi proteins and Piwi-interacting RNAs (piRNAs) are best known for their roles in suppressing transposons and promoting fertility. Yet piRNA biogenesis and its mechanisms of action differ widely between distantly related species. To better understand the evolution of piRNAs, we characterized the piRNA pathway in C. briggsae, a sibling species of the model organism C. elegans. Our analyses define 25,883 piRNA producing-loci in C. briggsae. piRNA sequences in C. briggsae are extremely divergent from their counterparts in C. elegans, yet both species adopt similar genomic organization and transcription program that drive piRNA expression. By examining production of Piwi-dependent secondary small RNAs, we identified a set of protein-coding genes that are evolutionarily conserved piRNA targets. In contrast to C. elegans, small RNAs mapped to ribosomal RNAs or histone transcripts are not hyper-accumulated in C. briggsae. Instead, we found that fewer introns in transcripts are associated with hyper-accumulation of small RNAs. Together our work highlights evolutionary conservation and divergence of the nematode piRNA pathway and provides insights into its role in endogenous gene regulation. Small RNA high-throughput sequencing was done using RNA from adult wild type and prg-1 mutant animals in C. elegans and C. briggsae. C. briggsae PRG-1 IP was preformed using adult animals. Differential gene expression analysis, and other analyses, was done to identify C. briggsae piRNA loci, as well as smRNA abundance changes between wild type and prg-1 mutants.

Piwi蛋白（Piwi proteins）与Piwi互作RNA（Piwi-interacting RNAs，piRNAs）最广为人知的功能为抑制转座子并维持生殖能力。然而，piRNA的生物发生及其作用机制在远缘物种间存在显著差异。为深入解析piRNA通路的进化历程，我们对模式生物秀丽隐杆线虫（Caenorhabditis elegans，C. elegans）的姊妹种——布里格斯隐杆线虫（Caenorhabditis briggsae，C. briggsae）中的piRNA通路开展了系统表征。本研究共鉴定出布里格斯隐杆线虫中25883个piRNA生成位点。尽管布里格斯隐杆线虫的piRNA序列与秀丽隐杆线虫的同源序列差异极大，但二者均采用相似的基因组组织模式与转录程序驱动piRNA的表达。通过检测Piwi依赖的次级小RNA的生成情况，我们鉴定出一组进化保守的piRNA靶蛋白编码基因。与秀丽隐杆线虫不同，布里格斯隐杆线虫中未出现靶向核糖体RNA或组蛋白转录本的小RNA过度积累现象；相反，我们发现转录本内含子较少的基因与小RNA的过度积累存在关联。综上，本研究揭示了线虫piRNA通路的进化保守性与分化特征，并为其在内源基因调控中的作用提供了新的见解。本研究分别提取秀丽隐杆线虫与布里格斯隐杆线虫的成年野生型及prg-1突变型动物的RNA，用于小RNA高通量测序；同时利用成年布里格斯隐杆线虫样本完成了PRG-1免疫沉淀（IP）实验。通过差异基因表达分析及其他相关分析，我们鉴定了布里格斯隐杆线虫的piRNA位点，并比较了野生型与prg-1突变型样本中小RNA的丰度差异。