Systems-Based Analysis of the Sarcocystis neurona Genome Identifies Pathways That Contribute to a Heteroxenous Life Cycle

Blazejewski, Tomasz; Nursimulu, Nirvana; Pszenny, Viviana; Dangoudoubiyam, Sriveny; Namasivayam, Sivaranjani; Chiasson, Melissa A.; Chessman, Kyle; Tonkin, Michelle; Swapna, Lakshmipuram S.; Hung, Stacy S.; Bridgers, Joshua; Ricklefs, Stacy M.; Boulanger, Martin J.; Dubey, Jitender P.; Porcella, Stephen F.; Kissinger, Jessica C.; Howe, Daniel K.; Grigg, Michael E.; Parkinson, John

Given similarities in gene numbers and exon lengths, we next determined the source of the additional sequence associated with To identify factors responsible for the increased intra-and intergenic region sizes, we performed a systematic analysis of repetitive regions across representative apicomplexans with the software tool RepeatModeler (20) . This analysis revealed that the S. neurona genome is rich in repeats largely associated with long interspersed nucleotide element (LINE) and DNA element sequences (class I and II transposons, respectively). Mapping of the repeats to scaffolds revealed that many of the repeats are associated with genes ( Fig. 1B) . Further comparisons of introns, exons, and intergenic regions showed clear differences in the repeat type based on the genomic context (Fig. 1C) . DNA element-type repeats were enriched in intronic regions and virtually absent from exons, suggesting evolutionary pressure against the integration of DNA elements within coding regions. Conversely, LINE-like repeats were equally distributed across exons, introns, and intergenic regions. In total, 31 Mbp of the S. neurona genome had repetitive sequences, compared to 17.9 Mbp of the E. tenella genome and 2.5 Mbp of the T. gondii genome ( Fig. 2A ). S. neurona displays a diverse set of repetitive elements. The repetitive sequences present in S. neurona are extraordinarily diverse, with 203 families of repeats discovered with RepeatModeler, compared to 101 families in E. tenella and 5 in Plasmodium falciparum. The majority of simple repeats within the S. neurona genome belong to more diverse families, unlike other apicomplexan parasites, where simple repeats are largely composed of short repeats (e.g., CAG n in E. tenella [21] ). For example, 33 simple repeat families in S. neurona were composed of consensus sequences with an average length of 287 bp. The average length of the simple repeats was 105 bp in S. neurona, compared to 48 and 68 bp in P. falciparum and E. tenella, respectively. Type II transposons, or DNA elements with 64,732 members, represent the largest family of repeats present in S. neurona, totaling 14.6 Mbp (11.5%) of the genome, considerably more than in E. tenella ( Fig. 2A) . All of the DNA elements identified belong to the "cut-and-paste" families of transposons, which propagate through genomes through excision and insertion of DNA intermediates. The most abundant family of DNA elements belonged to the CACTA-Mirage-Chapaev family of transposons, although a minority of Mutator-like elements was also identified. Active DNA transposons contain transposase genes; however, we were unable to detect any such gene within the S. neurona genome, suggesting that these DNA elements are ancient and degraded. Supporting this view, we found that the ratio of transversions to transitions in alignments of repetitive sequences to DNA repeat families was almost exactly 2:1, the statistically expected rate of mutation in the absence of evolutionary pressure.