Microbes never cease to surprise, as should be evident to long-time STC readers. Now we are presented with a novel mechanism that allows archaea to expand their gene content and metabolic capacity. This unforeseen discovery also illustrates the importance not only of having the right tools (and funding) but also of being prepared to find the unexpected.
In addition to being phylogenetically distinct from both bacteria and eukaryotes, some archaea have unique metabolic functions. One of these functions involves the production of methane by methanogens, as well its oxidation via the reverse reaction. These processes impact the cycling of methane, a potent greenhouse gas, and are therefore key to the current global climate crisis. These reactions, however, occur only under anaerobic conditions. Which is why Basem Al-Shayeb and colleagues working in Jill Banfield's lab at UC Berkeley set out to study samples collected from oxygen-depleted, anoxic locations where methane is produced and oxidized, such as groundwater, sediments and wetland soils, in the United States.
To study these environmental samples, the group focused on sequencing the metagenomic DNA, which refers to the collection of genomes found in a particular location. By carefully assembling and teasing out properties of the resulting DNA sequences they stumbled upon "enigmatic genetic elements" that they named Borgs. Borgs are, as you may recall, an alien civilization in the Star Trek saga that assimilates and transforms individuals to achieve perfection. In the case of archaea, Borgs are large extrachromosomal elements of unusual properties that also assimilate DNA from their hosts and expand their functional capacity. The Borg genomes are linear and large, ranging from approximately 600 kb to almost 1Mb in size. They contain unusual stretches of perfect tandem repeats (multiple adjacent copies of DNA) both within genes and between them (Figure 1), as well as regions involved in replication. These extrachromosomal elements, however, are distinct from any previously described autonomous genetic elements in archaea. Borgs are not like chromosomes because they lack ribosomal RNA genes; they are linear and much larger than known viruses and code for no known capsid proteins; and they lack the characteristic plasmid partitioning systems. So, if they're not chromosomes, viruses or plasmids, Borgs they are for now.
The initial metagenomic analysis of anoxic sites led to the assembly of four complete Borg genomes and one that was almost complete. These Borgs were given the colorful names of Black, Purple, Sky, Lilac, and Orange, respectively. As often occurs with sequenced environmental samples, most of the genes were novel, i.e., they had no similarity to genes in the reference database (Figure 2). Among those genes whose predicted proteins did have matches, the majority corresponded to archaeal proteins and specifically to Methanoperedens spp., archaea that are involved in methane oxidation. These extrachromosomal elements are therefore tightly associated with Methanoperedens, microbes that utilize an enzyme called MCR (methyl-CoM reductase) to break down methane.
Upon close examination of genes within one of these Borgs, the Lilac Borg, the authors found host-like MCR genes for methane oxidation. They also found genes for large multi-heme cytochromes (MHCs) involved in transfer of electrons to terminal electron acceptors during this process. The presence of these functions on Borg elements suggests that they increase the capacity of the host cell to oxidize methane (Figure 3). But of course, these large DNA elements also contain many other genes involved in multiple functions like energy metabolism, carbon utilization and stress responses, as well as possible defense systems.
Armed with information specific to Borg genomes, the next step was to see if these elements could be found in other locations. By examining metagenomic datasets from diverse environments, they identified 19 additional Borg sequences. Since then, six more complete genomes have been assembled and used to characterize the regions of their genomes that contain the tandem repeats. Interestingly, the repeat regions within genes do not disrupt the gene's open reading frame but rather form amino acid tandem repeats that are under strong selective pressure and contribute to the functional diversification of Methanoperedens spp.
Altogether the discovery of these unusual Borg extrachromosomal elements exemplifies the power of culture-independent methods to probe yet uncultured environmental microbes. It also required some exceptional sleuthing abilities to be able to discern the presence of novel DNA elements based solely on DNA sequence analysis. Thus, the arsenal of extrachromosomal elements in archaea is increased to include Borgs, which in addition to being involved in gene transfer also expand the metabolic activities of host cells.
Many of the hypotheses raised in this study, such as the novelty of these elements and their functional roles, must await experimental validation using cultured representatives, which have proven difficult to obtain. However, the ecological relevance of Borgs is impressive, as are the numerous contributions and cutting-edge work done in the Banfield lab. Thus, it was a with great delight that we learned at STC that Jill Banfield is the recipient of the of van Leeuwenhoek Medal 2023. More to come soon on this...