Barthelonids represent a deep-branching metamonad clade with mitochondrion-related organelles predicted to generate no ATP

Abstract

We here report the phylogenetic position of barthelonids, small anaerobic flagellates previously examined using light microscopy alone. Barthelona spp. were isolated from geographically distinct regions and we established five laboratory strains. Transcriptomic data generated from one Barthelona strain (PAP020) were used for large-scale, multi-gene phylogenetic (phylogenomic) analyses. Our analyses robustly placed strain PAP020 at the base of the Fornicata clade, indicating that barthelonids represent a deep-branching metamonad clade. Considering the anaerobic/microaerophilic nature of barthelonids and preliminary electron microscopy observations on strain PAP020, we suspected that barthelonids possess functionally and structurally reduced mitochondria (i.e. mitochondrion-related organelles or MROs). The metabolic pathways localized in the MRO of strain PAP020 were predicted based on its transcriptomic data and compared with those in the MROs of fornicates. We here propose that strain PAP020 is incapable of generating ATP in the MRO, as no mitochondrial/MRO enzymes involved in substrate-level phosphorylation were detected. Instead, we detected a putative cytosolic ATP-generating enzyme (acetyl-CoA synthetase), suggesting that strain PAP020 depends on ATP generated in the cytosol. We propose two separate losses of substrate-level phosphorylation from the MRO in the clade containing barthelonids and (other) fornicates.

Reduced mitochondria provide an essential function for the cytosolic methionine cycle

“…Leger et al (2017) predicted GCS and SHMT in the MROs of all free-living Fornicates and proposed that glycine degradation correlates with their lifestyle for an unclear reason. Moreover, GCS and SHMT were predicted to be localised in the MRO of other free-living Metamonada, such as Barthelona sp ., Anaeramoeba flamelloides and A. ignava (Stairs et al, 2021; Yazaki et al, 2020) and free-living Archamoebae Mastigamoeba balamuthii and Pelomyxa schiedtii (Nývltová et al, 2015; Záhonová et al, 2022). Our data are consistent with these observations and provide a potential reason for maintaining GCS and SHMT in these organelles – production of folates to supply cytosolic 1C metabolism.…”

Evidence for an Independent Hydrogenosome-to-Mitosome Transition in the CL3 Lineage of Fornicates

“…Each of these groups evolved from and contains free-living forms, but they are currently dominated by endobiotic taxa living in anoxic niches provided by animal bodies. Recently, two more metamonad lineages, represented by the poorly studied free-living barthelonids ( Barthelona and its formally undescribed relatives) and anaeramoebids (presently equal to the genus Anaeramoeba ), have been discovered ( Yazaki et al, 2020 ; Stairs et al, 2021 ). Each of these lineages, especially anaeramoebids, has been instrumental in illuminating the nature of the MRO ancestral for metamonads: this is inferred to have been a genome-lacking organelle possessing functions typical for the hydrogenosome, i.e., production of H 2 as a metabolic waste product related to the generation of ATP, but with additional physiological functions typical of the classical mitochondrion, such as a disulfide relay system, propionate production, and amino acid metabolism ( Stairs et al, 2021 ).…”

“…This vast time span has allowed for a lot of evolutionary experimentation with the ancestral metamonad MRO, resulting in a high diversity of MRO forms in extant metamonads ( Jerlström-Hultqvist et al, 2013 ; Zubáčová et al, 2013 ; Leger et al, 2017 ; Yazaki et al, 2020 ; Stairs et al, 2021 ). Thus, some members, including Anaeramoeba , Parabasalia, and some lineages of the Fornicata, have retained hydrogenosomes.…”

Phylogenomics invokes the clade housing Cryptista, Archaeplastida, and Microheliella maris

“…The analyses of small subunit ribosomal DNA (SSU rDNA)-one of the most popular gene markers for organismal phylogeny-succeeded in finding the phylogenetic homes of many eukaryotes/lineages, of which morphological information was insufficient to resolve their phylogenetic affiliations [15][16][17][18] . More recently, orphan eukaryotes/lineages, as well as newly found eukaryotes have been subjected to phylogenomic analyses 7,8,[19][20][21][22][23][24][25][26] .…”