Field of Science

Detecting mutual exclusivity of gene families

Gene networks are popularly used in systems biology to show functional associations among genes within a single genome, taking advantage of available experimental data on intermolecular interactions. Co-evolutionary networks are another way of showing functional associations among genes, in this case using presence/absence patterns of homologous genes across genomes to predict likely interaction partners. A nice example from proteins that I'm interested in are the components of the selenocysteine incorporation machinery for incorporating the amino acid selenocysteine into growing peptides. Not all organisms utilise selenonocysteine, but those who do encode a whole package of genes for its synthesis, charging onto tRNA and delivery to the ribosome. If a gene X was to be found in only that strange collection of (not always closely related) organisms with the selenocysteine machinery, chances are that X either uses selenocysteine or is also involved in its metabolism. As an aside, STRING is a really nice web application for visualising networks of functional associations compiled from various sources of evidence (co-occurence, co-expression, gene neighbourhood, and experiments).

A new paper by Zhang et al. in GBE presents a new and interesting approach for analysing co-evolutionary networks, by detecting Mutually Exclusive Orthologous Modules (MEOMs). In their words: "A MEOM is composed of two sets of gene families, each including gene families that tend to appear in the same organisms, such that the two sets tend to mutually exclude each other (if one set appears in a certain organism the second set does not)."

MEOMs are interesting because they reflect the replacement of one set of genes by another. This could be due to lineage-specific or environment-specific adaptations. The authors analyze a co-evolutionary network based on 383 organisms from across the tree of life and find that MEOMs most often include gene families involved in transport, energy production, metabolism, and translation.  They suggest that changes in the metabolic environment of an organism require adaptation to new sources of energy, and this triggers of replacement of genes, complexes and pathways in individual lineages. They also find many outer membrane proteins in their MEOMs, suggesting that as these proteins interact with the extracellular environment, they are frequently replaced during adaptation.

It's all very interesting, and I hope the authors will consider making a searchable web interface to their database of MEOMs. Their supplementary data is a bit awkward to navigate, and this kind of data is just crying out for visualisation.  I would love to be able to scan proteins in my data sets for potential MEOM membership.

Xiuwei Zhang, Martin Kupiec, Uri Gophna, & Tamir Tuller (2011). Analysis of Co-evolving Gene Families Using Mutually Exclusive Orthologous Modules Genome Biology and Evolution : 10.1093/gbe/evr030

The ancestral ribosome: my reservations

I work on deep evolution of ribosome-associated proteins, so of course I'm very much excited by research on deep evolution of ribosomal RNA. However, I have some concerns about some of the work in this field relating to the composition and structure of the ancestral ribosome, or as sometimes called, proto-ribosome. Actually, it’s less that I have concerns about the work, more that I have some small, but (at least to me) important concerns about the interpretations and subsequent speculations. Anyway, the other day, as I listed to Ada Yonath's and Loren Williamson's talks at the Suddath Symposium on the Ribosome, I was reminded about these concerns, and decided it's probably a good idea to blog about them. 

Before I start moaning, I want to stress that it's really exciting that people are trying to answer such deep evolutionary questions, and I genuinely think they have made some interesting and important discoveries about the relative ages of parts of the ribosome, and about small catalytic RNAs that can behave like ribosomes, I just don't think those catalytic RNAs are ancestral ribosomes. I think they are perhaps some shared component of ancestral and modern ribosomes

What it comes down to is that in general the people working on proto-ribosomes are assuming that evolution proceeds from small and simple to large and complex. In fact, this is not necessarily the case, as I have blogged about previously. Small and perfectly formed is hard to evolve, while big and clumsy with time for optimisation is less hard.

Figure 1. My crude drawing to demonstrate evolutionary progression. Green is the modern ribosome, with the predicted ancient parts in red. Black is sequence nonhomologous to the modern ribosome. A) shows the idea that is often presented: a small protoribosome gets bigger over time. B) shows my hypothesis: continual loss and gain of sequence with some retention of a conserved core.
Ada Yonath’s talk at the symposium on the ancestral peptidyl transferase centre (PTC, the region where peptide bonds are formed between amino acids) really captured my imagination. The PTC is buried right in the middle of the ribosome and consists of  two fragments of rRNA with rotational structural symmetry between the P (peptidyl) site tRNA binding rRNA and the A (acceptor) tRNA binding rRNA. This symmetrical region is highly conserved in sequence (98% identity among organisms), but not between each symmetrical unit. Ada proposes that this symmetrical region is the oldest part of the ribosome, and that this minimal region is a functional machine on its own. In support of this, the CCA-end of tRNA fits in perfectly, and their structural studies indicate it could provide a rotary motion of tRNAs that is required get peptidyl transfer. This is a really nice story, and so far I’m totally in support. What I have problems accepting is that this minimal rRNA dimer IS all that was present of the protoribosome (as in fig 1A). Why could there not have been extra RNA around it that was replaced during evolution (as in fig 1B)? Via the online participation (which was fantastic by the way) for the symposium I asked Ada about this:

Gem: The small symmetrical region might be the only region modern ribosomes have in common with the ancestral proto-ribosome. But it almost seems TOO streamlined. Could the protoribosome actually have been bigger than that core region, and there could have been loss as well as gain of sequence? 

Ada: we haven’t thought of that… but you can speculate anything.

That’s exactly my concern, that you can speculate anything in this field. There are very few clues to go on, and they don’t give anything conclusive. Where evidence dries up, all you can do are thought experiments, based on examples we know of.  And we know from extant ribosomes that there have been lineage-specific loss and gain of sequence.  Good examples are mitochondrial ribosomes which have lost a good deal of rRNA and replaced it with protein.

A similar model of ribosome evolution to Ada's, proposing progressive addition of rRNA onto a minimal but functional PTC frame is presented by Bokov and Steinberg, Nature (2009). In this paper, the authors examine the inter-domain interactions and structural dependencies in the large subunit to infer relative age. Again, this is great, fascinating work, and it is also consistent with a model of replacement and optimisation, rather that the “aggrandizement” that they presume in their model.

After Ada Yonath’s talk in the symposium came Loren Williams, who also works on figuring out the ancestral ribosome. Williams and colleagues compared the sequences and structures of  archaeon H. marismortui and bacterium T. thermophilus ribosomes and found that sequence and conformational similarity of the rRNAs are greatest near the PTC, and diverge smoothly with distance from it.  They show a beautiful figure of the ribosome as an onion, which makes their point perfectly.

Again, these particular results are very clear and interesting, it’s just some of the assumptions about the evolutionary process that I have issues with. I noticed in the talk that Loren consistently equated “conserved” with “old.” In fact, “conserved” usually means “important”. Jamie Williamson who was in the audience also made this point during the talk, and Loren replied that he could not argue with that. In the case of the ribosome, the central parts are not only involved in catalysis, they are also important for maintaining the three dimensional structure. So they are very important. It’s the same reason why proteins show strong conservation of buried amino acids.

Some other evolutionary statements and suppositions by Loren also were a bit iffy, such as: "mitochondrial ribosomes are running evolution backwards." Yikes. Drastically cutting down rRNA and replacing with protein independently in multiple lineages is definitely not running evolution backwards… in fact evolution is never, ever backwards. I also have a problem with supposing things that it isn’t necessary to suppose: Loren hypothesises that the ribosome binding tails of ribosomal proteins are older than the globular domains, and were originally non-coded, they then became fused to globular domains. There really is no evidence for this as far as I can see. The tails and insertions that protrude into the ribosome are very biased in amino acid content, and if they’re anything like ribosome-binding extensions of translation factors such as IF3, they readily appear and vary in length and primary sequence during evolution.  These sort of structures seem easy to add.

Maybe my complaints can be considered to be petty in a field that is necessarily rife with speculations, but I just think it’s important not to push the speculations too far, in order to keep our scientific integrity and not become like the cranks that publish their “evolutionary biology” in the Journal of Cosmology. For example, I loved the first half of Ada’s talk, but she finished it with a discussion of the ability of her two symmetrical fragments to dimerise, and suggested that in a population of these fragments, their non-uniform tendency to dimerise was a kind of "pre darwinian Darwinian” ribosome evolution that took place in the prebiotic world. She also suggested that these fragments may also be proto-tRNAs. For me, this is too far removed from the evidence, and these are speculations too far.

BUT! Having said all that, wild speculation is bloody well fun, so I will offer my own hypothesis (see fig. 1 B). I think the first ribosome could have been big, flabby and clumsy, an amalgamation of RNAs that were perhaps already involved in some other catalysis such as nucleic acid polymerisation, that through chance flopping around, managed to catalyse (probably in a very inefficient way) peptide bond formation. The efficiency of bond formation between particular amino acids may have been influenced by the certain nucleic acids being polymerised in the active site, as in some primitive ‘code’. This protein synthesising proto-machine maybe had nothing recognisably in common with modern ribosomes, but it was subsequently fine-tuned through loss of gain of sequence until it became something resembling the ribosome that we know and love. 

Refs and further reading

Belousoff MJ, Davidovich C, Zimmerman E, Caspi Y, Wekselman I, Rozenszajn L, Shapira T, Sade-Falk O, Taha L, Bashan A, Weiss MS, & Yonath A (2010). Ancient machinery embedded in the contemporary ribosome. Biochemical Society transactions, 38 (2), 422-7 PMID: 20298195

Bokov K, & Steinberg SV (2009). A hierarchical model for evolution of 23S ribosomal RNA. Nature, 457 (7232), 977-80 PMID: 19225518

Hsiao C, Mohan S, Kalahar BK, & Williams LD (2009). Peeling the onion: ribosomes are ancient molecular fossils. Molecular biology and evolution, 26 (11), 2415-25 PMID: 19628620