Age-related macular degeneration (AMD) is a leading cause of blindness in humans, and the leading cause of visual impairment during advanced age. The condition comes in two basic forms, the most severe of which is untreatable. Called geographic atrophy (GA), this condition involves the steady destruction of the retinal pigment epithelium, a layer of tissue in the eye that is essential for the health and maintenance of the photoreceptors in the retina. Loss of the pigment epithelium means certain death for the photoreceptors, and that means visual impairment and then blindness for the affected person.
A major publication in Nature last month (Kaneko et al., "DICER1 deficit induces Alu RNA toxicity in age-related macular degeneration," Nature 17 March 2011) now points to one likely cause of AMD, and in the process provides a chilling example of what can happen when the parasitic Alu elements in our genomes (see the previous post for an introduction) are left unrestrained.
The story begins with experiments that showed that a very interesting enzyme, DICER1 (which we'll just call Dicer) was markedly depleted in the retinas of people with the GA form of AMD. Dicer (as its name is meant to convey) specializes in chopping things up. Specifically, it chops up microRNAs, which are small (of course) pieces of RNA that cells make. This is a more precise and important job than it seems: Dicer carefully trims the microRNA into a version that is fully active in the control of gene expression. The resulting pieces of RNA are tiny (21 letters long) but potent, able to substantially reduce the expression of the genes they target. (The phenomenon is called RNA interference and its discovery revolutionized cell biology by giving biologists a simple way to manipulate gene expression.) The authors were probably examining Dicer in the context of AMD because several previous reports had shown that loss of Dicer led to problems in the development of the retina.
So, having found that Dicer was reduced in diseased retinas, the authors showed that this deficit can lead to the disease process. (The mere correlation they started with need not mean that the Dicer problem was causative in any way.) They genetically engineered mice that lacked Dicer in their retinas, and the mice got a nasty GA. And so, after only the first illustration of the article, the biologists had strong evidence that depletion of Dicer could lead to AMD, and that alone is a significant finding.
But why Dicer and AMD? The first and most reasonable hypothesis was that the loss of Dicer led to a failure to trim microRNAs and thus to an overall problem with the microRNA-based gene control system. To test this hypothesis, the authors deleted gene after gene in the microRNA processing system (Dicer is one of several enzymes in that system) and failed to see any retinal problems in any of the resulting mice. The surprising conclusion: the problem that leads to AMD when Dicer is depleted is not a problem with microRNA processing. Dicer's critical role lies elsewhere.
But where? Well, Dicer specializes in chopping up RNA, and specific types of RNA (double-stranded RNA, to be exact). So the authors looked first to see if there was excess double-stranded RNA in retinas of people with GA. Sure enough, there was a big difference between diseased and normal retinas. So they did some nifty biochemistry to grab those excess double-stranded RNAs and identify them. And something jumped out: they were getting Alu RNAs. Click.
Recall that Alu elements are the most abundant type of repeated DNA sequence in the human genome. The human genome contains more than a million of these things, and they account for more than 10% of the genome by themselves. They are mobile genetic elements, meaning that they love to hop from place to place in the genome, and they do this by making RNA copies of themselves. This means that we can expect, at least sometimes, to see Alu RNAs in cells.
Back to the article. What followed was a series of compelling experiments that showed that Alu RNAs are specifically enriched in the doomed cells of the RPE of diseased retinas, and that Dicer does indeed destroy Alu RNAs. The critical next step is an experiment you've probably already identified: to see whether adding Alu RNA to normal cells can kill them. It can. First the authors showed that generic Alu elements can kill the RPE cells when artificially introduced. Then they did something really cool: they took one of the Alu elements that they had fished out of a diseased human retina and introduced it into a mouse retina. This is what they saw (Figure 4c):
On the left are the retinas that got the Alu element. The devastation is most apparent in the red pictures in the bottom row, which show the outlines of happy normal cells on the right and a nightmare of degeneration on the left. Alu elements, when left unchecked, destroy the retinal pigment epithelium and lead to AMD.
But there's one more experiment, a coup de grace, that would really nail this. Here's how it goes. Dicer depletion leads to degeneration and to AMD. Check. It also leads to increased Alu element RNA, and that leads to degeneration and to AMD. Check and check. Now, if those observations add up to a coherent explanation, then we can make the following prediction: the degenerative effect of Dicer depletion should be negated by erasing the runaway Alu elements. The authors took their engineered mouse, which normally gets nasty GA, and then depleted the Alu elements using a simple technique that mimics RNA interference. In short, they induced AMD by depleting Dicer, then attempted to prevent the AMD by killing the Alu elements. And it worked (Figure 5c).
Focus again on the red pictures in the bottom row. Both retinas depicted are lacking Dicer. The trashed retina on the right got no help; the one on the left was rescued by the erasure of the most widely-expressed Alu elements.
The implications of this work are very significant. For one thing, the authors have identified a target for further work aimed at reversing or preventing AMD. But we've also learned something important about Alu elements. As we might expect while considering a parasite that is the most abundant mobile genetic element in the human genome, Alu elements do not tend to have our best interests in mind, and thus their activity must be regulated, and even opposed. We already knew that they can wreak havoc by jumping indiscriminately or by destabilizing genome structure; now Kaneko et al. have shown us another dark side of the Alu world. They write:
This also is, to our knowledge, the first example of how Alu could cause a human disease via direct RNA cytotoxicity rather than by inducing chromosomal DNA rearrangements or insertional mutagenesis through retrotransposition, which have been implicated in diseases such as α-thalassaemia, colon cancer, hypercholesterolemia, and neurofibromatosis.And ominously, they point out that it's possible that other experiments (and diseases) in which Dicer is lost or depleted may be explained by Alu toxicity rather than by problems in microRNA processing. Until now, biologists didn't know just how precious Dicer was.
In the next post I'll conclude by discussing these findings in light of various creationist claims on the topic, including an ongoing series at Reasons To Believe.
Kaneko, H., Dridi, S., Tarallo, V., Gelfand, B., Fowler, B., Cho, W., Kleinman, M., Ponicsan, S., Hauswirth, W., Chiodo, V., Karikó, K., Yoo, J., Lee, D., Hadziahmetovic, M., Song, Y., Misra, S., Chaudhuri, G., Buaas, F., Braun, R., Hinton, D., Zhang, Q., Grossniklaus, H., Provis, J., Madigan, M., Milam, A., Justice, N., Albuquerque, R., Blandford, A., Bogdanovich, S., Hirano, Y., Witta, J., Fuchs, E., Littman, D., Ambati, B., Rudin, C., Chong, M., Provost, P., Kugel, J., Goodrich, J., Dunaief, J., Baffi, J., & Ambati, J. (2011). DICER1 deficit induces Alu RNA toxicity in age-related macular degeneration. Nature, 471 (7338), 325-330 DOI: 10.1038/nature09830