Comparative genomics reveals insight into the evolutionary origin of massively scrambled genomes

Abstract

Ciliates are microbial eukaryotes that undergo extensive programmed genome rearrangement, a natural genome editing process that converts long germline chromosomes into smaller gene-rich somatic chromosomes. Three well-studied ciliates include Oxytricha trifallax, Tetrahymena thermophila and Paramecium tetraurelia, but only the Oxytricha lineage has a massively scrambled genome, whose assembly during development requires hundreds of thousands of precise programmed DNA joining events, representing the most complex genome dynamics of any known organism. Here we study the emergence of such complex genomes by examining the origin and evolution of discontinuous and scrambled genes in the Oxytricha lineage. This study compares six genomes from three species, the germline and somatic genomes for Euplotes woodruffi, Tetmemena sp., and the model ciliate Oxytricha trifallax. To complement existing data, we sequenced, assembled and annotated the germline and somatic genomes of Euplotes woodruffi, which provides an outgroup, and the germline genome of Tetmemena sp.. We find that the germline genome of Tetmemena is as massively scrambled and interrupted as Oxytricha's : 13.6% of its gene loci require programmed translocations and/or inversions, with some genes requiring hundreds of precise gene editing events during development. This study revealed that the earlier-diverged spirotrich, E. woodruffi, also has a scrambled genome, but only roughly half as many loci (7.3%) are scrambled. Furthermore, its scrambled genes are less complex, together supporting the position of Euplotes as a possible evolutionary intermediate in this lineage, in the process of accumulating complex evolutionary genome rearrangements, all of which require extensive repair to assemble functional coding regions. Comparative analysis also reveals that scrambled loci are often associated with local duplications, supporting a gradual model for the origin of complex, scrambled genomes via many small events of DNA duplication and decay.

Exploration of the Nuclear Proteomes in the Ciliate Oxytricha trifallax

“…This also helps prevent cells which have finished digesting their food from entering the starvation-induced encystment stage of their life cycle, which is resistant to the lysis buffer used in the nuclear isolation procedure. While this method was originally developed for DNA isolation and typically includes a secondary size-based separation afterwards [ 27 , 28 ], nuclear proteins do not share the same size bias between the MAC and MIC that chromosomes do. Despite this lack of secondary enrichment, MAC and MIC samples are noticeably different when separated on an SDS-PAGE gel, as well as when blotted with an anti-H3 histone antibody.…”

Nuclear dualism without extensive DNA elimination in the ciliateLoxodes magnus

“…57 Old MACs in Loxodes have higher and more variable DNA content than recently matured MACs, 38 but this may be nonspecific amplification in senescent nuclei, as we did not observe distinct subclusters of MACs by DNA content (Figure 1D) nor evidence for differential amplification. Chromosome breakage and unscrambling were not directly addressed here, but unscrambling has only been found in conjunction with IES elimination, 5,58 and is likely also absent. To assess chromosome breakage, Loxodes telomeres, which have thus far eluded our detection, will need to be identified.…”

“…43% vs. 4% of the human genome, 62 it is surprising that we have not detected compelling DNA transposon homologs in Loxodes, as they are numerous in all ciliate MIC genomes examined to date (Figure 4A). 5,6,8,9,58 This may simply reflect the most recent wave of mobile element proliferation in this particular strain, or higher deleteriousness of DNA transposons than retrotransposons.…”

From germline genome to highly fragmented somatic genome: genome-wide DNA rearrangement during the sexual process in ciliated protists

“…Transposable elements were detected as described in other ciliates (Chen and Landweber 2016 ; Feng et al 2022 ). Briefly, representative Oxytricha TBE ORFs (Genbank accession AAB42034.1, AAB42016.1, and AAB42018.1) were used as queries to search TBEs in the S .…”

“…Even for species in the same class, the patterns of PGR can be quite different. For example, the somatic genomes of the spirotrich genus Euplotes (e.g., E. vannus and E. woodruffi ) have similar size and structure to, but larger MDSs than, O. trifallax (Chen et al 2014 , 2019 ; Feng et al 2022 ). Furthermore, only about 4–7% of the genes in the germline genome are scrambled in Euplotes , far lower than the 20–30% seen in Oxytricha .…”