Where do genes come from? The case of the “green” parasites

This essay was written by Don Williamson and was first published in the 2001 Mill Hill Essays.

Recent advances in molecular biology frequently make headlines in the media for their possible role in leading to “breakthroughs” in the diagnosis or treatment of genetic diseases or cancer. Outside of laboratories, however, it is less commonly realised that one of the most striking consequences of present day molecular technology is our ability to explore in detail the genetic make-up of thousands of creatures – plants, protozoa, fungi and animals, including of course, ourselves. This means firstly that we can establish the genetic relatedness of different organisms and classify them in a meaningful way that does not rely on subjective assessments of behaviour or appearance. Secondly, and more importantly, it lets us reconstruct the evolutionary pathways that have led to the appearance of all species of animals or plants living around us today. As a consequence, we are increasingly able to ask a fundamental question; where did genes come from? Sometimes the answer is totally unexpected.

Of course the immediate source of your genes is obvious; they came from your parents, and they got them from their parents, and so on. And if you were a single-celled organism like a bacterium or an amoeba, you would have acquired them from the parent cell that was you before it (you) split in two. This suggests that in the natural world the genome of any given species is totally segregated from that of any other, and for sexually reproducing metazoan organisms like ourselves there is some truth in this. However, we now know that most vertebrate species, for instance, carry chromosomal genes derived from viruses and bacteria, and there can be few ‘higher’ organisms (animals, plants, protozoa and fungi) that are free of genes acquired in ancient times from totally unrelated species. Indeed, it is becoming ever more apparent that as the evolutionary web of life was unfolding, it was greatly moulded by such inter-specific transfer of genes, in the form of bits of DNA, between different organisms. Increasingly our sophisticated modern tools for DNA analysis are letting us stumble across examples of these long past events, known in the trade as “lateral (or horizontal) transfer”. As just one example, it has now been shown that as much as 18% of the genome of the E. coli bacterium was acquired by lateral transfer from other organisms.

The grand-daddy of lateral transfers is the key evolutionary event which led to possession by almost all so-called ‘higher’ organisms of mitochondria. These are the little intracellular bodies (organelles) that provide energy by carrying out the biochemical changes involved in respiration, and which almost all organisms made up of higher cells depend on. It seems certain now that these vital little structures, which retain a few of their original genes on their own piece of DNA, originated as bacteria (even perhaps a single bacterium, the evidence pointing to a single event) which in the distant past were (or was) swallowed up by a primitive (but unidentified) ancestor. Technically this event is referred to as endosymbiosis, and the captive organism, while maturing fully into its new role, may be called an endosymbiont.

The endosymbiotic origin of mitochondria is an idea with a long history, but it is only recently that modern techniques have let us recognise that the genes carried on the mitochondrion’s own little genome are indeed of bacterial origin. However, these genes are only the runt of the large raft of genes that came in with the original symbiont, many of which must have been essential to its well-being. It now seems that as part of the endosymbiotic deal, many of these genes must eventually have been transferred to the host’s nucleus and integrated into its chromosomes. To be sure, there are conceptual difficulties with this idea; after all, genes have to be tightly regulated, and proteins destined for particular intracellular compartments need to be kitted out with attachments (signal sequences) that enable them to be correctly addressed, but there is little doubt the chromosomes of cells like our own do carry genes recognisably bacterial in origin.

The endosymbiosis leading to mitochondria was certainly a key event in fashioning the present-day biological world. Arguably however it is overshadowed by the even more important one which led to the evolutionary emergence of plants and algae. In addition to mitochondria, green organisms all carry photosynthetic organelles known as chloroplasts (or plastids), so named because of the chlorophyll they contain. Like the mitochondria these also arose as a result of an ancient endosymbiotic event, but this time the ingested victim came from the ranks of organisms which used to be called “blue-green algae” but are actually bacteria and are now known as cyanobacteria.

These bacteria had developed, on their own account, the ability to acquire energy from sunlight by photosynthesis, using various pigmented compounds for the purpose. It is now known that chloroplasts of all photosynthetic higher organisms originally evolved, more or less directly, from a progenitor cell swallowing a cyanobacterium. As with the mitochondria, many of the genes required for maintenance of the new victim were transferred to the capturing cell’s nucleus, and although less essential ones were presumably lost, somehow the system developed so that the acquired organelle benefited the host, and for this reason has been retained for millions of years of evolution. In many ways, the development of chloroplasts has to have been the most important event in the evolution of life, underpinning as it does the overall ecosystem that acquires energy from the sun, on which we all ultimately depend.

The reality of these endosymbiotic events is attested by two lines of evidence. Firstly, analysis of the nucleotide sequences of the genes carried on the DNA retained by mitochondria and chloroplasts shows them to be very similar to the corresponding genes of the free-living micro-organisms believed to have been the donors. Secondly, we can now spot the bacterial origin of many of the genes transferred to the nucleus. But in any case, endosymbiosis is not just a theoretical concept thought up to explain the origins of the organelle genes. There are plenty of examples, especially among protozoa, where it is often relatively easy to recognise bacterial symbionts microscopically. In many of these cases the symbiosis may be one-way only, and removal of the symbiont has little effect on the host organism. However, in one of the best known examples, a stable two-way endosymbiosis was seen to happen quite quickly. In some astonishing experiments , the American researcher K. W. Jeon and his colleagues in Buffalo, USA, witnessed an endosymbiotic integration actually occurring and maturing to full-scale interdependence of host and symbiont in the laboratory. For many years Jeon had been maintaining the giant amoeba A. Proteus with no problems. One year it was noticed that the organisms became sick, growing more slowly and dividing less often. Nevertheless they continued to grow and were carefully nurtured by Jeon’s team. Right from the start it was realised that the apparent cause of their sickness was the appearance in their cytoplasms of numerous bacteria. Clearly the amoebae had found the initial invasion disagreeable, but after a few months, their growth improved, albeit not quite to the same level as before. By this time the bacteria had become extremely numerous in the cytoplasm, to the extent of around 40,000 per amoeba. They still looked recognisably like bacteria. The astonishing thing was that now the amoebae were totally dependent on the little invaders; exposure to antibiotics of the kind that normally only affect bacteria resulted in the death of the amoebae themselves. Obviously something in the nature of the biochemical interactions between the host and the invaders had changed. The host amoebae were now completely dependent on the activities of the bacterial remnants but, at the same time, the bacteria had clearly lost their independence; they could easily be transfected into different amoebae, but could no longer grow in the outside world.

Some evolutionary endosymbioses leading to photosynthesis have been more complicated than the simple event described above. Two groups of algae dignified unforgettably as ‘chlorarachniophytes’ or ‘cryptomonads’ have acquired a very unusual chloroplast by their ancestors having ingested not just a cyanobacterium but a whole eukaryotic alga, itself already equipped with a “conventional” chloroplast. This constitutes a whole different ball game, and is referred to as secondary endosymbiosis. The algal chloroplast resulting from the primary endosymbiotic event had already gone through the process of shedding unwanted bacterial genes and transferring only those needed for its own maintenance to the nucleus of the algal host. It is reasonable to suppose that when the secondary endosymbiosis took place, these vital genes would be held on to and retained somewhere in the new (secondary) host. By comparison with conventional chloroplasts one would have expected them to have been moved to the secondary nucleus. However, the chlorarachniophytes and cryptomonads have muddied the waters because, after millions of years of evolution, they have retained, trapped in a complex chloroplast bounded by 4 membranes rather than the usual 2, what appears to be a remnant of the ingested algal nucleus. This unique body is known as a nucleomorph, and its precise location between plastid membranes 2 and 3 (see the accompanying figure) speaks eloquently of the evolutionary history of the chloroplast itself.

The nucleomorph has three small and compactly organised chromosomes, and the genome of one species has been completely sequenced. It carries hundreds of functional genes, about thirty of which are specifically required for the enclosing plastid, but many more are regarded as ‘housekeeping’ genes, needed for their expression. The organisation, evolutionary origin and activities of the nucleomorph are hot topics for research in several laboratories, but it is at least clear that although genes for a few of the chloroplast’s essential maintenance tools have wound up in the final host’s nucleus, many more are still retained on nucleomorph chromosomes. This is not too surprising, since it hardly seems likely that in the long term, energy would have been devoted to maintaining the complex structure of the little organelle, with its own chromosomes, membrane and means of gene expression, unless it did something useful. Short of a totally unrelated (and unexpected) function, retention of chloroplast-related genes would seem as good a role as any. Nevertheless it is not too fanciful to think that the nucleomorph-containing organisms are just “frozen” intermediates in the evolution of chloroplasts from secondary endosymbiosis.

There is in fact good reason to suppose that secondary endosymbiosis can go the extra mile, generating a more or less conventional residual chloroplast without the complication of an accompanying nucleomorph. In this case, the chloroplast of the victim is retained, in a heavily trimmed down state, and all the victim’s nuclear-encoded genes needed to maintain what’s left of it are also retained, but unlike the chlorarachniophytes and cryptomonads, they are all transferred to the new host’s nucleus. At the same time all the victim’s unwanted genes and all morphological traces of its nucleus are discarded. Evidence for this comes from a totally unexpected quarter, and one that, surprisingly, has important medical implications for humans. It has resulted from study of the malaria parasite and its relatives.

The malaria parasites belong to a huge group of microbial parasites, a number of which cause serious disease in man and animals. Known as “Apicomplexa” they all are obliged to grow and reproduce inside the cells of higher organisms and none of them is photosynthetic. Nevertheless, despite this, many of them were recently found to possess the residue of a chloroplast carrying, like all organelles of this ilk, their own little piece of DNA. The apicomplexans are thought to have evolved, billions of years ago, from free-living micro-organisms known as dinoflagellates. These rather aggressive little creatures are abundant in the present day sea, where they are sometimes responsible for the “toxic blooms” that afflict our shores in summer weather, but they have a very long evolutionary history. Many of them are photosynthetic, having acquired the plastids they need for this by long ago swallowing cyanobacteria. It now appears however, that billions of years ago one of them (or possibly even an earlier progenitor) excelled its own voracious capabilities by swallowing not just a cyanobacterium but, like the cryptomonads, a fully-fledged alga, itself already equipped of course with a chloroplast. This greedy microbial pirate then, like Jeon’s amoebae, grew to like some of the ingested plastid’s activities, and selectively retained the basic structure of the organelle together with these preferred functions, while disposing of all the unwanted nuclear apparatus like chromosomes, membranes and nuclear division apparatus. Of course it is easy to write about this in a couple of sentences, but the reality must have been incredibly complex. As with all long-term endosymbioses, the retained structures, be they plastids, mitochondria or even nucleomorphs, required the retention of a host of genes needed to maintain and operate the physical structures involved. And, as already mentioned, these genes must have been regulated so that they only made the right amounts of their products at the right times, and they would have had to acquire tags so that the cell’s apparatus would know where to send the proteins they specified. All this must have taken unimaginable years, although Jeon’s experiments with amoebae suggested that some sort of accommodation between the host and its newly acquired booty can sometimes be achieved very rapidly. In any event however, billions of years on from the initial piracy, the strange little chimeric creature we have been discussing eventually gave rise to apicomplexans like the malaria parasite.

The mind boggles at the complexity of this process, but there is undeniable evidence in the case of the malaria parasite (and related apicomplexans) that the nucleus is studded with genes required for activities thought to be solely the province of the residual chloroplast. Notice that word “thought”. The truth is that at the moment we do not know exactly what the plastid’s function actually is. It is certainly not photosynthesis, as the parasites never see daylight, but apart from this energy-gaining activity, normal fully functional chloroplasts in plants undertake synthesis of metabolites for the host cell, lipids, amino-acids and the like – bits and pieces required for assembly of all manner of cell apparatuses and biochemical activities. It is likely that the residual plastid genes have been retained for one of these functions, though it is still a big puzzle why all the genes involved have not been moved into the nucleus. Present evidence suggests the synthesis of lipids or membranes may be the remaining key role, but this research is ongoing and at the moment everything is up for grabs. All we know for sure is that the malarial plastid organelle has some essential function, because drugs that selectively knock it out are lethal for the host cell.

Herein lies a practical footnote to what otherwise might be seen as a somewhat abstruse academic quirk of interest only to ivory-towered researchers. Most biochemical pathways, particularly the “house keeping” ones, are very similar in all organisms. This poses a big problem for designers of anti-parasitic drugs, since even if a compound is found that can access a target enzyme in a parasite, it may also target the same enzyme in the host, and therefore be toxic. This is true for instance, even for anti-mitochondrial drugs, because cells of all creatures have mitochondria. However, we do not carry even the remains of plastids in our bodies, and our genes are not (for the most part) of plant-like origin, so genes inherited by the malaria parasites as a result of their ancestors’ long ago piratical gobbling of little green plants – the algae – have little chance of close resemblance to those of their host animals – us. There is every possibility therefore that drugs which do act against specific plastid-related enzymes in the parasites may be without effect on the host.

Magic bullets! I hear you cry, and you are right. There is now a world-wide “industry” seeking out malarial plastid-related genes in the hope of identifying potential targets, and I cannot resist pointing out that the original identification of the plastid DNA was made in our laboratories in Mill Hill. But we must be careful. It would be quite wrong to herald these advances in a journalistic fashion as a much hyped “breakthrough”. There are hundreds of plastid-related genes carried by the nucleus, and detailed and painstaking work is needed to identify them. Even with the help of the nearly completed sequencing of the malaria parasite’s entire genetic blueprint, it will take a long time to find new effective drugs to combat the little beasts. But at least we now have an excitingly logical avenue to follow, and all because of our modern ability to find out where genes come from.

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