Malaria has confounded some of the best minds of the past century. A hundred years after the discovery that mosquitoes transmit Plasmodium falciparum, the major parasite that causes human malaria, we still do not know enough about the disease to defeat it permanently. But the papers on pages 498–542 of this issue1, 2, 3, 4, 5, 6, 7, describing the complete genome sequence of P. falciparum, may eventually lead to new drugs and vaccines, and will certainly be an invaluable guide to future research. These papers are a testament to the success of a six-year project undertaken by an international consortium of labs and funding agencies.
Why genome sequencing?
First, a bit of background. The malaria parasite leads a complicated life (Fig. 1), existing mainly inside liver cells and red blood cells in its human host and, when residing in mosquitoes (notably Anopheles gambiae), being associated with the insect's gut and salivary glands. It undergoes several transformations along the way. The stages of its life cycle were originally described more than 100 years ago and were given names based on morphology, such as merozoite, trophozoite and gametocyte (in humans), and zygote, ookinete and sporozoite (in mosquitoes). One of the most curious features of the human stages is the human immune response — there is much immune activity, but this does not control the infection effectively, nor afford protection against future infections.
Figure 1: Life cycle of the parasite Plasmodium falciparum.
a, When a parasite-infected mosquito feeds on a human, it injects the parasites in their sporozoite form. These travel to the liver, where they develop through several stages, finally producing merozoites which invade and multiply, via the trophozoite stage, in red blood cells. Eventually, up to 10% of all red cells become infected. (Clinical features of malaria, including fever and chills, anaemia and cerebral malaria, are all associated with infected red blood cells, and most current drugs target this stage of the life cycle.) The merozoites in a subset of infected red blood cells then develop into gametocytes. b, When another mosquito bites the infected human, it takes up blood containing gametocytes, which develop into male and female reproductive cells (gametes). These fuse in the insect's gut to form a zygote. The zygote in turn develops into the ookinete, which crosses the wall of the gut and forms a sporozoite-filled oocyst. When the oocyst bursts, the sporozoites move to the mosquito's salivary glands, and the process begins again.
High resolution image and legend (46K)Despite massive efforts to eradicate the disease in the 1950s and early 1960s, more people are infected with malaria in Africa today than at any other time in history. Over 500 million people are infected with the disease worldwide, and one-quarter of the population is at risk of infection. More than a million children die of malaria each year, mostly in Africa. And those individuals who survive suffer a combination of anaemia and immune suppression that leaves them vulnerable to other fatal illnesses. Alarmingly, drug resistance in the parasite is now widespread.
These stark facts emphasize the need to find new treatments for the disease and new ways of preventing it. The genome project described in this issue1, 2, 3, 4, 5, 6, 7 was conceived with these goals in mind. With the wealth of information now available at the click of a mouse, malaria researchers have an unprecedented opportunity to find genes that are potentially unique to, or at least substantially different in, P. falciparum compared with other species; such genes may make good drug targets, with less risk of side effects.
Even before the whole genome had been sequenced, new drug targets were being identified from searches of the partially assembled sequence data for unique genes8. But the total sequence will provide a more complete picture of the parasite's inner workings and the chance to identify vulnerable aspects. So just what have we learnt about the parasite's biology from this package of papers, which comprises its genome sequence1, 4, 5, 6; a comparison of its genome with that of a rodent malaria parasite, P. yoelii yoelii2; and two proteomics studies of the proteins expressed at different stages in the parasite's life cycle3, 7? Where are the potential weaknesses? And what have we discovered about the parasite's means of evading the human immune response?
Metabolism
One notable feature of the parasite's genome1 is the apparent absence of genes for proteins that, in other species, are key to metabolism and the energetics of mitochondria — cellular powerhouses, which produce the energy-storing molecule ATP. For example, the consortium found no predicted genes for two protein components of ATP synthase, a mitochondrial ATP-producing enzyme. (At present, many of the genes are only 'predicted': they have been identified by gene-searching algorithms, but have not yet been confirmed as bona fide genes.) Similarly, there are apparently no genes for components of a conventional NADH dehydrogenase complex, another key mitochondrial enzyme. Perhaps P. falciparum generates and stores energy by using novel proteins or mechanisms — potential drug targets. That the mitochondria are active, at least in sporozoites and gametocytes, seems likely, given that the proteomics analyses3, 7 detected fragments of enzymes involved in some typical mitochondrial processes, including the tricarboxylic-acid cycle and oxidative phosphorylation.
Also interesting is the number of predicted genes — some 10% — that encode proteins associated with the apicoplast1. This essential cellular compartment is known to be important for the biosynthesis of fatty acids and isoprenoids, components of many membrane proteins, and for iron metabolism. But analysis of these genes should reveal other possible functions, and so new drug targets. The genome sequence also identifies the molecules within the apicoplast that are the targets of several existing drugs9.
The complex life cycle of P. falciparum means that the parasite has had to adapt to several different environments. So it is also intriguing that, compared with the genome of the free-living budding yeast, the parasite genome1 encodes a limited number of predicted transporter proteins for the active uptake of nutrients from the environment. In fact, entire classes of transporters seem to be missing. It may be that several genes in this class have been overlooked because they are made up of many small coding regions, which can be missed by gene-prediction algorithms. But, taken at face value, this surprising finding implies that adequate amounts of nutrients recognized by the transporters must be present at all stages of the parasite life cycle, so that there is no selective advantage in having many transporters with differing substrate specificities. Alternatively, the parasite may use previously identified pores or channels to acquire nutrients10, 11.
Regulating protein levels
During its life cycle, P. falciparum undergoes several developmental changes. One of the most dramatic is sexual differentiation and the formation of gametes, male and female reproductive cells. The proteomics studies3, 7 of these stages have coincidentally shed light on a fundamental question: how does the parasite regulate the levels of its proteins? The genome1 encodes relatively few predicted proteins that control the transcription of genes into messenger RNAs (the first step in making a protein). Moreover, there seem to be few transcriptional regulatory elements in the genome — or at least, there are few elements that are known from other organisms. Yet the proteomics analyses and previous studies show that protein abundance is tightly regulated.
The proteomics studies also show that proteins involved in processing mRNAs and in protein synthesis (translation) are expressed at higher levels in gametocytes, particularly female gametocytes, than in other stages. Interestingly, proteins that are present in early zygotes — which are produced from gametocytes — seem to be absent in gametocytes, although the mRNAs encoding these proteins are abundantly present. All of this is consistent with the proposal12 that the regulation of protein levels is controlled through mRNA processing and translation, rather than by gene transcription. Perhaps this is a general feature of the parasite — another potential drug target.
In addition, one of the proteomics studies3 reveals groups of genes whose regulation appears to be coordinated. Some simultaneously expressed genes are clustered in the genome; comparison of these genes and their flanking sequences may provide further insight into how they are regulated.
Immune evasion
Arguably the most striking features of the P. falciparum genome are the regions near the ends of each chromosome1. This is where families of genes that encode surface proteins, such as the var genes, are found. These proteins, or antigens, can sometimes be recognized by and thus stimulate the human immune system. But they have a great capacity for change, which occurs partly through the exchange of material between chromosome ends. As the genome sequence shows, the very ends of the chromosomes — the telomeres — have a complex arrangement of sequences that may facilitate such exchange (as described in ref. 13) and thereby lead to immune evasion.
The general structure of the chromosome ends is similar to that in the rodent parasite P. yoelii yoelii2. But, surprisingly, the genes that encode the variant surface antigens in P. falciparum are not found in P. yoelii yoelii, which has a different family of variant genes, originally described in a less virulent human parasite, P. vivax14. This is interesting, because it suggests that P. yoelii yoelii, which is often used as a model of P. falciparum, is in some respects more similar to P. vivax. It is tempting to speculate that, despite their dissimilar sequences, the genes at the ends of the P. falciparum and P. yoelii yoelii chromosomes have similar functions. But that remains to be seen.
Finally, research on the P. falciparum var genes has focused on their role in enabling infected red blood cells to stick to small blood vessels in the brain. This feature is associated with the fatal form of the disease, cerebral malaria. So it is interesting that one of the proteomics analyses3 reveals that the peptides derived from many of the var genes occur in sporozoites, which are produced in mosquitoes and invade the human liver during the initial infection. These results point to possible alternative functions for var gene products.
The complete picture
One of the most exciting aspects of this huge undertaking is that it can be related to other work. We now have the genome of the mosquito A. gambiae15, together with draft sequences of the human genome16, 17, and so can get a better handle on the interactions among three species that have long been evolving together. It is well known that certain variations in human genes are associated with a reduced susceptibility to malaria, and analysis of different human populations will no doubt reveal more on this. A close look at the mosquito genome should provide similar insights. Study of the parasite genome will reveal much about how P. falciparum interacts with its host and carrier, and more about the genes involved in parasite recognition by the human immune system. Decoding the information in these genomes, and translating it into effective remedies, is both a challenge and an opportunity for the scientific community.
