Abstract
ABSTRACTSeparate germline and somatic genomes are found in numerous lineages across the eukaryotic tree of life, often separated into distinct tissues (e.g., in plants, animals, and fungi) or distinct nuclei sharing a common cytoplasm (e.g., in ciliates and some foraminifera). In ciliates, germline-limited (i.e., micronuclear-specific) DNA is eliminated during the development of a new somatic (i.e., macronuclear) genome in a process that is tightly linked to large-scale genome rearrangements, such as deletions and reordering of protein-coding sequences. Most studies of germline genome architecture in ciliates have focused on the model ciliates Oxytricha trifallax, Paramecium tetraurelia, and Tetrahymena thermophila, for which the complete germline genome sequences are known. Outside of these model taxa, only a few dozen germline loci have been characterized from a limited number of cultivable species, which is likely due to difficulties in obtaining sufficient quantities of “purified” germline DNA in these taxa. Combining single-cell transcriptomics and genomics, we have overcome these limitations and provide the first insights into the structure of the germline genome of the ciliate Chilodonella uncinata, a member of the understudied class Phyllopharyngea. Our analyses reveal the following: (i) large gene families contain a disproportionate number of genes from scrambled germline loci; (ii) germline-soma boundaries in the germline genome are demarcated by substantial shifts in GC content; (iii) single-cell omics techniques provide large-scale quality germline genome data with limited effort, at least for ciliates with extensively fragmented somatic genomes. Our approach provides an efficient means to understand better the evolution of genome rearrangements between germline and soma in ciliates.
Highlights
Separate germline and somatic genomes are found in numerous lineages across the eukaryotic tree of life, often separated into distinct tissues or distinct nuclei sharing a common cytoplasm
We describe how the germline genome architecture is associated with gene family size; in C. uncinata, the largest gene families, which appear Chilodonella specific, are enriched with scrambled genes
To explore the germline genome architecture of Chilodonella uncinata, we compared the characterization of germline sequences derived from a “traditional” gel-based method to those from a single-cell omics approach
Summary
Separate germline and somatic genomes are found in numerous lineages across the eukaryotic tree of life, often separated into distinct tissues (e.g., in plants, animals, and fungi) or distinct nuclei sharing a common cytoplasm (e.g., in ciliates and some foraminifera). In Chilodonella uncinata (in the class Phyllopharyngea), the germline genome is composed of more “traditional” chromosomes [11,12,13] while the somatic chromosomes are present as “gene-sized” nanochromosomes that are maintained at variable copy numbers As a result, this ciliate, described as having an extensively fragmented genome, has a somatic nucleus that harbors Ͼ20 million nanochromsomes [14,15,16]. Because of difficulties in culturing and the high level of amplification of somatic genomes compared to the germline (which contributes to contamination in germline DNA preps), traditional methods for sequencing germline-limited DNA are fairly laborious and costly in terms of time and benchwork This has led to limitations in the phylogenetic breadth of explorations of ciliate germline genomes to a few model species for which cultures can provide sufficient numbers of cells (often in the millions) and for which time-tested germline isolation techniques exist. The limitations on the ability to extract quality germline micronuclear DNA with sufficient yields for highthroughput library construction, especially considering the loss of DNA associated with each manipulation and purification step [17], has likely been the greatest barrier to studies of germline genomes in non-model ciliates
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