Minimal genomes of mycoplasma-related endobacteria are plastic and contain host-derived genes for sustained life within Glomeromycota

Abstract

Arbuscular mycorrhizal fungi (AMF, Glomeromycota) colonize roots of the majority of terrestrial plants. They provide essential minerals to their plant hosts and receive photosynthates in return. All major lineages of AMF harbor endobacteria classified as Mollicutes, and known as mycoplasma-related endobacteria (MRE). Except for their substantial intrahost genetic diversity and ability to transmit vertically, virtually nothing is known about the life history of these endobacteria. To understand MRE biology, we sequenced metagenomes of three MRE populations, each associated with divergent AMF hosts. We found that each AMF species harbored a genetically distinct group of MRE. Despite vertical transmission, all MRE populations showed extensive chromosomal rearrangements, which we attributed to genetic recombination, activity of mobile elements, and a history of plectroviral invasion. The MRE genomes are characterized by a highly reduced gene content, indicating metabolic dependence on the fungal host, with the mechanism of energy production remaining unclear. Several MRE genes encode proteins with domains involved in protein-protein interactions with eukaryotic hosts. In addition, the MRE genomes harbor genes horizontally acquired from AMF. Some of these genes encode small ubiquitin-like modifier (SUMO) proteases specific to the SUMOylation systems of eukaryotes, which MRE likely use to manipulate their fungal host. The extent of MRE genome plasticity and reduction, along with the large number of horizontally acquired host genes, suggests a high degree of adaptation to the fungal host. These features, together with the ubiquity of the MRE-Glomeromycota associations, emphasize the significance of MRE in the biology of Glomeromycota.

Keywords: genome contraction; genome plasticity; horizontal gene transfer; vertical transmission.

Fig. 1.

16S rRNA phylogeny reveals inter- and intrahost diversity of MRE populations. Cloned 16S rRNA gene sequences were obtained from three individual spores (A, B, or C) representing populations MRE-RC, MRE-RV, and MRE-CE (bolded). Numbers following spore designations indicate individual sequences. Reference MRE represent operational taxonomic units defined at a 97% sequence similarity level. Bayesian posterior probabilities greater than 0.90 are shown above branches. Branches with maximum likelihood bootstrap support over 70% are thickened.

Fig. 2.

Contig structures reveal that MRE genomes are highly recombinant, resulting in disruptions of gene synteny. Contigs are shown as black elongated ovals, and colored lines indicate sequences and gene order. Sequences that are identical and have the same gene order are depicted in the same color. The bases corresponding to identity between the contigs are indicated. (A) Example of two contigs of MRE-RV sharing identical sequences (orange) followed by completely different genetic content (green and red). (B) Example of three contigs of MRE-RC sharing regions of identity (pink) whereas flanking regions contain unrelated gene content (green, white, yellow, orange, and blue).

Fig. 3.

Horizontal gene transfer of the SUMO protease gene from AMF to MRE. Homologs of the AMF SUMO protease-encoding gene (ESA14994/KI282387) are present in the MRE-RV and MRE-CE metagenomes. The gene from MRE-RV (GenBank locus tag MRERV_24c013) is shown here as a representation of the horizontal gene transfer and subsequent evolution of the gene. Note that the AMF SUMO protease sequence used is that of R. irregularis (Ri) because it is the only AMF sequence available; this sequence may not be the original one that was transferred. The full DNA sequence of the SUMO protease gene of AMF (Ri) is shown as a gray bar. The corresponding primary transcript sequence is shown below, with exons (E) represented by blue bars, and introns (In) shown as pale blue bars. The DNA sequence of the gene from MRE-RV is displayed as red bars, with the horizontal line indicating loss of sequences/deletions in the MRE genome. The MRE gene still maintains traces of the introns.

Fig. 4.

Multigene phylogeny reveals shared ancestry of MRE and the members of the Mycoplasmataceae family. Phylogenetic reconstructions were performed on the concatenated amino acid sequences of the following genes: dnaG, infC, nusA, rplA, rplB, rplC, rplE, rplF, rplM, rplN, rplP, rplT, rpmA, rpsB, rpsC, rpsE, rpsJ, rpsS, and smpB. Bayesian posterior probabilities greater that 0.90 are shown above branches. Branches with maximum likelihood bootstrap support over 70% are thickened. Data from MRE-CE were not included due to high diversity of 16S rRNA gene sequences in this population (Fig. 1).