Rampant nuclear insertion of mtDNA across diverse lineages within Orthoptera (Insecta)

Abstract

Nuclear mitochondrial pseudogenes (numts) are non-functional fragments of mtDNA inserted into the nuclear genome. Numts are prevalent across eukaryotes and a positive correlation is known to exist between the number of numts and the genome size. Most numt surveys have relied on model organisms with fully sequenced nuclear genomes, but such analyses have limited utilities for making a generalization about the patterns of numt accumulation for any given clade. Among insects, the order Orthoptera is known to have the largest nuclear genome and it is also reported to include several species with a large number of numts. In this study, we use Orthoptera as a case study to document the diversity and abundance of numts by generating numts of three mitochondrial loci across 28 orthopteran families, representing the phylogenetic diversity of the order. We discover that numts are rampant in all lineages, but there is no discernable and consistent pattern of numt accumulation among different lineages. Likewise, we do not find any evidence that a certain mitochondrial gene is more prone to nuclear insertion than others. We also find that numt insertion must have occurred continuously and frequently throughout the diversification of Orthoptera. Although most numts are the result of recent nuclear insertion, we find evidence of very ancient numt insertion shared by highly divergent families dating back to the Jurassic period. Finally, we discuss several factors contributing to the extreme prevalence of numts in Orthoptera and highlight the importance of exploring the utility of numts in evolutionary studies.

Figure 1. Numts of three mitochondrial genes (COI, COII, ND5) are extremely abundant across the phylogenetic diversity within Orthoptera.

The y-axis shows the proportion of numts from the total number of clones generated based on PCR using conserved primers. If nearing 1, most clones generated are numts. If nearing 0, most clones generated are orthologous mtDNA.

Figure 2. Characteristics of numts as measured by base composition and uncorrected p-distance from orthologs.

A: COI-like numts; B: COII-like numts; C: ND5-like numts.

Figure 3. Co-analysis of numts and mtDNA using COI-like numts of Stenopelmatus fuscus (Ensifera: Stenopelmatidae).

Grey terminals represent orthologous COI across Orthoptera. The maximum likelihood analysis recovered a monophyletic group consisting of COI-like numts and the ortholog from S. fuscus. Paleonumts are denoted by yellow with red outline, and neonumts are denoted by yellow with blue outline. Those numts with premature stop codons, insertions, and deletions, are indicated by orange, green, and blue squares, respectively. Numbers on nodes indicate bootstrap support values.