Endosymbiosis of Frankia cluster-2
Actinobacteria of the genus Frankia engage in root endosymbiosis with actinorhizal plants belonging to the Fagales, Cucurbitales and Rosales. The genus Frankia consists of four different clades. Strains belonging to cluster-1, -2, and -3 exhibit host-specificity which plants they can nodulate, while cluster-4 strains do not engage in symbiosis. Cluster-2 is the earliest divergent clade, and most of its members could not be cultured thus far, with the exception of Frankia coriariae BMG5.1 and BMG5.30. Unlike other nodule symbioses where ammonia is the nitrogen source exported to the host, it has been shown in nodules of Datisca glomerata that an assimilated nitrogen metabolite, namely arginine, is exported to the host. In the study on bacterial transcription patterns during symbiosis presented here, it is suggested that in nodules of Ceanothus thyrsiflorus an assimilated nitrogen source, most likely asparagine, is delivered by Frankia cluster-2. Analysis of the carbon metabolism indicates that citrate could be the carboxylate delivered from the host to the bacteria. Analysis of the tricarboxylic acid cycle indicates that the glyoxylate shunt is missing in genomes of Frankia cluster-2. During symbiosis the pathway mostly works linear between input of carbon skeletons and the output of assimilated nitrogen metabolites. This feature could explain why Frankia cluster-2 strains have a low saprotrophic potential.
Analysis of genomes of Frankia cluster-2 have shown that murC is present in two gene copies within the genome of cluster-2 and cluster-3 strains, but only present as one copy in cluster-1 and cluster-4. MurC plays a key role in biosynthesis of peptidoglycan, a polymer that makes up a thick layer as part of cell wall of Gram-positive bacteria. Phylogenetic analysis show one of the two copies encodes a protein highly similar to MurC protein of other actinobacteria, and therefore is considered the ancestral copy, murC1. The other copy, murC2, can be found in all Frankia clusters and encodes a protein showing more similarity to MurC from cyanobacteria and plants. This similarity could be explained through either horizontal gene transfer, which is not unlikely as Frankia can be found intracellularly in its host plants. The other possible scenario is that a gene duplication event was followed by convergent evolution, leading to similarity between MurC2 and the MurC proteins of cyanobacteria and plants. This would allow for plasticity in the structure of the peptidoglycan layer of the cell wall.
The features of nitrogen and carbon metabolism, as well as the phylogeny of MurC, indicate Frankia cluster-2 has undergone several adaptations to intracellular endosymbiosis.
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