Genomes on the edge: programmed genome instability in ciliates

Abstract

Ciliates are an ancient and diverse group of microbial eukaryotes that have emerged as powerful models for RNA-mediated epigenetic inheritance. They possess extensive sets of both tiny and long noncoding RNAs that, together with a suite of proteins that includes transposases, orchestrate a broad cascade of genome rearrangements during somatic nuclear development. This Review emphasizes three important themes: the remarkable role of RNA in shaping genome structure, recent discoveries that unify many deeply diverged ciliate genetic systems, and a surprising evolutionary "sign change" in the role of small RNAs between major species groups.

Figure 1. Simplified Ciliate Life Cycles

(A and H) Reproductive, vegetative growth occurs a sexually by cell division, including mitosis of the germline micronucleus (MIC, indicated by a circle) and amitosis of the somatic macronucleus (MAC, indicated by an oval). (B) Starvation induces conjugation between compatible mating types, initiating the nonreproductive sexual cycle. (C) Meiosis of the MIC produces haploid gametic nuclei. (D and E) (D) Exchange of haploid micronuclei and fertilization produces two new, diploid, zygotic nuclei (E). (F) Mitosis of zygotic nuclei produces two identical micronuclei, and one nucleus begins to differentiate into a new MAC. (G and H) (G) Degradation of the old MAC occurs during differentiation of the new MAC. Mature cells (H) enter again into vegetative growth (Nowacki et al., 2009).

Figure 2. Cladogram of Ciliate Genera Discussed in the Text, along with Other Representative Eukaryotic Genera

The branching order, branch lengths, and approximate scale bar, for reference, are based on Parfrey et al. (2011), with the addition of Euplotes based on Chang et al. (2005).

Figure 3. Structural Elements of IES Regions and Transposon-Derived Genes or Systems

Magenta boxes, macronuclear-destined sequence (MDS); light green, internal eliminated sequences (IES) or transposable elements. Inverted light green arrowheads indicate inverted repeats at ends of deleted sequences. Dark green triangles indicate short direct repeats (pointer sequences). Blue bars indicate open reading frames (ORFs). Solid thin arrows show correspondence between DNA sequences in the MIC and MAC, and black boxes indicate telomeres on MAC chromosomes. (A) Shown on the left is a schematic scrambled gene in Oxytricha or Stylonychia. In Oxytricha, thousands of MIC-encoded Tc1/mariner transposases (orange) likely participate in removal of IESs (indicated by dotted arrows) as well as themselves (Nowacki et al., 2009). (B) In oligohymenophorean ciliates, represented by Paramecium, IES excision requires a domesticated PiggyBac transposase encoded in the MAC. Paramecium has only TA dinucleotides as pointers and also flanking Tc1/mariner transposons in the MIC (Arnaiz et al., 2012). Euplotes crassus, a spirotrich, similarly has nonscrambled IESs with TA dinucleotides flanking both IESs and Tec transposons (Jacobs and Klobutcher, 1996). Tetrahymena differs from Paramecium in having mostly imprecise excision of longer intergenic IESs, with some precisely excised IESs flanked by TTAA repeats. (C) Schematic illustration of eukaryotic nonautonomous and autonomous DNA transposons. The autonomous elements (right) encode transposases that can mobilize truncated and simplified nonautonomous elements (left) throughout the genome. The structure of these elements mirrors the structure of ciliate IESs (nonautonomous elements) and their controlling transposons (autonomous elements), most likely reflecting their evolutionary origins and development (see Klobutcher and Herrick, 1997).

Figure 4. Two Models of RNA-Mediated Genome Rearrangements in Ciliates

(A) In Tetrahymena, bidirectional transcription of the MIC genome produces double-stranded RNA, which is processed into scnRNA duplexes. The Piwi protein Twi1p loads scnRNAs in the cytoplasm and transports them into the parental MAC to scan the somatic transcriptome. This process enriches for scnRNAs that do not pair with homologous sequences from the parental MAC. The MIC-limited scnRNAs are then transported into the developing MAC, where they recognize and mark IES regionsonMIC chromosomes. IES excision and telomere addition produce mature MAC chromosomes. In both panels, magenta rectangles denote MDSs, and combs indicate sRNA. (B) In Oxytricha, transcription of either strand of gene-sized chromosomes in the parental MAC produces telomere-containing template RNAs (blue numbered line) during conjugation. Either these template RNAs or other long noncoding RNAs are processed into 27 nt piRNAs. These form a complex with the Piwi protein Otiwi1 that transports them into the newly developing MAC, where the piRNAs recognize and mark the MDS portions of the MIC chromosome that are retained during genome rearrangement. The maternal template RNAs are also transported to the developing MAC, where they guide correct MDS ordering of numbered segments 1–6 (including inversion of segment 5) and DNA repair at recombination junctions to produce mature somatic chromosomes that are capped with short telomeres (small vertical black bars).