Identification and analysis of functional associations among natural eukaryotic genome editing components

Abstract

During development in the ciliate Paramecium, excess DNA interspersed throughout the germline genome is deleted to generate a new somatic genome. In this process, most of the intervening DNA is excised by a Piggybac-derived transposase, assisted by small RNAs (scnRNAs and iesRNAs) and chromatin remodelling. As the list of genes involved in DNA elimination has been growing, a need for a general approach to discover functional relationships among these genes now exists. We show that deep sequencing-based comparisons of experimentally-induced DNA retention provide a sensitive, quantitative approach to identify and analyze functional associations among genes involved in native genome editing. This reveals two functional molecular groups: (i) iesRNAs/scnRNAs, the putative Piwi- and RNA-binding Nowa1/2 proteins, and the transcription elongation factor TFIIS4; and (ii) PtCAF1 and Ezl1, two proteins involved in chromatin remodelling. Comparative analyses of silencing effects upon the largely unstudied regions comprising most developmentally eliminated DNA in Paramecium suggests a continuum between precise and imprecise DNA elimination. These findings show there is now a way forward to systematically elucidate the main components of natural eukaryotic genome editing systems.