Complete Mitochondrial Genomes of Ancyromonads Provide Clues for the Gene Content and Genome Structures of Ancestral Mitochondria

Abstract

Mitochondria of eukaryotic cells are direct descendants of an endosymbiotic bacterium related to Alphaproteobacteria. These organelles retain their own genomes, which are highly reduced and divergent when compared to those of their bacterial relatives. To better understand the trajectory of mitochondrial genome evolution from the last eukaryotic common ancestor (LECA) to extant species, mitochondrial genome sequences from phylogenetically diverse lineages of eukaryotes-particularly protists-are essential. For this reason, we focused on the mitochondrial genomes of Ancyromonadida, an independent and understudied protist lineage in the eukaryote tree of life. Here we report the mitochondrial genomes from three Ancyromonadida: Ancyromonas sigmoides, Nutomonas longa, and Fabomonas tropica. Our analyses reveal that these mitochondrial genomes are circularly mapping molecules with inverted repeats that carry genes. This inverted repeat structure has been observed in other mitochondrial genomes but is patchily distributed over the tree of eukaryotes. Ancyromonad mitochondrial genomes possess several protein-coding genes, which have not been detected from any other mitochondrial genomes of eukaryotes sequenced to date, thereby extending the known mitochondrial gene repertoire of ancestral eukaryotes, including LECA. These findings significantly expand our understanding of mitochondrial genome diversity across eukaryotes, shedding light on the early phases of mitochondrial genome evolution.

Keywords: Ancyromonas; Fabomonas; Nutomonas; rps5; AlphaFold; Diphoda; Foldseek; LECA; Opimoda; inverted repeats.

FIGURE 1

Complete maps of four Ancyromonadida species mitochondrial genomes. Coding regions are colored based on their function (see color key in figure). Black bold arrows indicate inverted repeats. Each map was drawn by OGDRAW and edited manually.

FIGURE 2

Conservation of synteny and gene repertoire of inverted repeats. Conservation of synteny around ribosomal genes (A) and complex I genes (B) is highlighted in a blue band. tRNA genes are omitted in the synteny analysis. (C) Gene repertoire of inverted repeats.

FIGURE 3

Structural comparison of Rps5 and Rpl10. (A) The N‐terminal (1–77 amino acid residues) and C‐terminal (119–198 amino acid residues) regions of Nutomonas longa Rps5 are shown in magenta and blue, respectively. The left and right in green show the experimentally determined structure of Escherichia coli Rps5 (chain E in pdb_00006nqb) and Homo sapiens nuclear‐encoded mitochondrial Rps5 (126–430 amino acid residues of chain AD in pdb_00003j9m), respectively. (B) Fabomonas tropica NYK3C Rpl10 is shown in magenta. The left and right in green show the experimentally determined structure of E. coli Rpl10 (chain I in pdb_00008upo) and H. sapiens nuclear‐encoded mitochondrial Rpl10 (77–197 amino acid residues of chain I in pdb_00003j9m), respectively. Structures were aligned by TM‐align.

FIGURE 4

Predicted 3D structure and TM helices of Nutomonas longa Orf1249. The structure of N. longa Orf1249 predicted by AlphaFold3 is shown in the front view and bottom view on the left and right, respectively. Transmembrane (TM) helices predicted by three in silico tools are shown in blue. TM helices predicted by at least one of three tools are shown in cyan.

FIGURE 5

Repertoire of protein‐coding genes in mitochondrial genomes across the eukaryote tree of life. Presence and absence of corresponding genes of various eukaryotes is shown by closed and open boxes, respectively. Genes of the mitochondrial genomes newly sequenced in this study are highlighted in dark blue and the presence of rps5 is indicated with the red box. The figure is updated based on several previous studies (Eglit et al. ; Kamikawa et al. ; Moreira et al. ; Yazaki et al. 2022). Phylogenetic relationships of eukaryotes are based on Williamson et al. (2025). The predicted gene contents of the last common ancestor (LCA) of Opimoda+, LCA of Ancyromonadida, and LCA of Eukaryota are shown at bottom, and numbers in parentheses are the total numbers of genes in each ancestral mitochondrial genome. Abbreviations of phyla: Am, Amoebozoa; An, Ancyromonadida; Ar, Archaeplastida; Cm, CRuMs; Di, Discoba; Ma, Malawimonadida; Op, Opisthokonta; Pa, Pancryptista; Pr, Provora; Sa, SAR.