A Worm's Bloody Eye March 3, 2000

I want to thank everyone who sent me comments on last week's article. I got more e-mails about that commentary than all the other ones combined. It is nice to know that some people actually DO read what I write. So, since I love getting responses, I'll continue my discussion on some recent investigations that may help shed light on how organisms change. This week I will discuss a novel 'eye' a worm uses to see.

In order for natural selection to have any effect on an organism, there must be variability. If every organism is exactly like every other one, there are no differences between them and no selective advantage can be gained by either of them. Now, we can see how mutations created by replication errors could create allellic differences in the genome. This provides an easy model for small differences between animals of the same species but how do new species come about? We are beginning to gain some insights into this process. One is that a structure that already exists can be used for a novel purpose or undergo a simple alteration, presenting new opportunities for selection to occur.

The observation that current uses for a structure are not the best indication of its initial utility is a constant theme in evolution. For example, it is difficult to understand how something like the lens of an eye could develop. Only the final product seems useful. An eye without a fully functional lens would not be able to see. However, we are looking at the endpoint of only one possible approach to sight. We observe that the clear lens allows us to see and marvel at the complex structure. But, that is backwards. We did not evolve sight because we have a clear lens. The possession of a clear lens simply gives us the ability to see in the manner that we do. A different sort of lens could very well have provided us a different mode of seeing.

Natural selection often co-opts structures or molecules that were developed for one reason, and uses them for another. These changes can then open up entirely new environmental niches for the organism to exploit. I'll discuss a recent publication examining the multiple purposes a protein can be put to and how the substitution of one protein for another may have had a striking effect on the life cycle of an organism.

The main constituents of the lens of vertebrate eyes are a family of proteins called crystallins. However, these proteins are also found in a wide variety of cells and are used for a host of processes, such as metabolic stress. In these settings, it is the enzymatic activity of the crystallins that is important, not their structural properties. In other cells they are expressed at relatively low levels. Only in the developing eye are they expressed at very high levels. Only in the eye are crystallins used to create a lens. Why only in the lens?

Well, the purpose of the lens is to refract light so that it can be focused on a specific area of the eye, allowing nerves to be stimulated, helping the organism get an idea of its surroundings. The protein must also be stable at high concentrations. So, there are structural considerations that are important, not enzymatic ones. Crystallin happened to be used because it has these properties, but perhaps it is possible that any protein that had these properties would have led to something like the vertebrate eye. Hard to "see", if we only look at the highly evolved, complex system we have today. Let's look at something similar, yet much simpler, and see whether there are any lessons.

Although crystallin homologues are found in Drosophila compound eyes, this is still a pretty complex system. There is a really simple one, though, found in worms or nematodes. Nematodes do not have eyes as we know them. However, several species have something that is similar. A bundle of nerves at the anterior of the worm serve to tell it where light is coming from. A small dark spot, or ocelli, made of the protein melanin lays over this bundle of nerves. It "shadows" the nerves when light is shined on it. The worm orients itself by rotating until the spot creates the greatest shadow over the nerves and then moves away from the light. This is called negative phototaxis. A very simple 'eye'. It reacts to light or shadow. But we know melanin is used for many other purposes. It is used here because it is opaque at high concentrations. While a transparent protein like crystallin would not be very useful in this setting, what about some other protein? What would happen if another protein, with different physical properties, was used? What effect would that have on the organism? Well, it appears that Nature may have provided us with an opportunity to examine these questions.

The nematode, Mermis nigrescens, is a parasite of grasshoppers. The host ingests eggs found on grass and the larvae grow inside up to a length of 10 cm. Late stage larvae break out of the host and burrow into the soil, where the adult will mature. In contrast to similar nematodes, this species does not always move away from light. Several years later, a pregnant female will emerge into the light and find a site on the grass to lay eggs. So these nematodes require a mechanism that allow them to discriminate between light levels, not the simple approach using melanin. The melanin system is binary, on or off. If a worm using this system displayed positive phototaxis and moved towards light, it would most likely die since nematodes can not withstand much exposure to direct sunlight. Perhaps the nematodes could develop some method to protect themselves from direct sun but there may be a simpler approach.

Instead of using an opaque protein, how about using one that absorbs some wavelengths of light but lets others through? Then, instead of only moving towards or away from direct light, the worm could move towards regions that are intermediate, such as the shadows in the grass. Perhaps the simple substitution of a protein that absorbs some light but is not completely opaque would be enough. That is what Mermis nigrescens appears to have done.

Here is a picture of a gravid female of Mermis. There is a large red pigmented area covering the nerves that can easily be seen. Electron micrographs display large, hexagonally-arrayed crystals in the cytoplasm of the cells. Only pregnant females displaying this red spot show positive phototaxis. What other protein do we know of that can be found in cells at high concentrations and is blood-red? Could it be hemoglobin?

Turns out that is exactly what the worms use. The worms show positive phototaxis over the same wavelengths of light at which hemoglobin absorbs. Amino acid sequencing of the purified protein from the ocelli indicated that the red pigment wasvery similar to a hemoglobin. The 'eye' hemoglobin sequences were used to clone the nematode cDNA via PCR. The protein is definitely a hemoglobin, having several amino acids which are absolutely conserved in all known hemoglobins. The levels of hemoglobin found in these cells is very similar to that seen in human red blood cells. Interestingly, the Mermis nigrescens hemoglobin has several amino acid residues that result in extremely high oxygen affinities. This makes sense since, in this setting, it would be critical to maintain the 'red' color of oxy-hemoglobin rather than the 'blue' color of deoxy-hemoglobin. And, since the protein does not need to ever give up its oxygen, as regular hemoglobin does, these affinities can be very large.

So, could a simple exchange in the expression of a protein be sufficient to generate a new species? Well, for two species to diverge, there must be little gene flow between the 2 species. A large change in reproductive strategies, such as laying eggs on blades of grass instead of in the soil, might accomplish this. Maybe a mutation that caused increased expression of a normal nematode hemoglobin in the ocelli, instead of melanin, was a critical step in opening up a new environmental niche for the worms to fill. A relatively simple change could have very large ramifications.

This hypothesis should be testable. Use genetic manipulation to switch the expression of hemoglobin for that of melanin in the shadowing spot of a species of nematode. What effect does this change have on the worms? What phototaxis is observed? Or take Mermis nigrescens and have it express melanin in the ocelli rather than the hemaglobin? These would be fascinating experiments.

Most organisms do what they can in order to reproduce. If they are born with a mutation that allows them to exploit a new environment, than they probably will exploit it. In this case, a relatively simple change in protein expression might have been the catalyst for the divergence of two species. The world has some pretty amazing things living on it, all waiting to move into new niches if the chance appears. I have talked about bacteria that can probably stand an atomic blast. But a worm that uses blood to see has to be one of the marvels of the world.