12 Oct 2009

MicroRNAs, tiny snippets of nucleic acid that wield far-reaching power in modifying gene expression, have taken a place in the panoply of factors that influence neurodegenerative disease. Many scientists think this budding field holds promise for understanding and, eventually, targeting these diseases therapeutically in fresh ways (see ARF related news story and Live Discussion).

Geneticist Stephen Cohen fell into the study of microRNAs when he happened upon an intriguing mutant. After showing that a particular microRNA, miR-8, is linked to an apparent neurodegenerative condition in fruit flies (Karres et al., 2007), he is now casting a wide net to discover any and all microRNAs that influence behavior and other phenotypes in Drosophila.

Born a Canadian, Cohen moved to Singapore in 2007 to become executive director of the Temasek Life Science Laboratory. (Meaning “sea town” in Javanese, the word Temasek refers to Singapore’s origins as a fishing village.) Prior to that, he bounced between North America and Europe several times, with stints at the European Molecular Biology Laboratory in Heidelberg, Germany; Baylor College of Medicine in Dallas, Texas; the Max Planck Institute in Tübingen, Germany; MIT; and the University of Toronto. ARF spoke with Cohen about life as a scientist in Singapore, his work on microRNAs, and his expectations for the future of the field.

Q&A

ARF: What drew you to Singapore?

SC: Curiosity about life in Asia combined with a good job opportunity. Scientists are very lucky that we can move our jobs around the world with us.

The life science community here is really exciting. The Singapore government has decided to invest in life sciences as a part of their national strategy. The view is that a knowledge-based economy is of strategic importance for the future. In fact, having no natural resources, it really is the obvious strategy. In a really short period of time, they have built up world-class institutes and attracted top talent from all over the world.

ARF: How does the culture of science here compare to other places you have worked?

SC: There are some regional differences, but they are subtle. The presence of a lot of people trained in Europe and North America greatly influences the culture of science here. The presence of a lot of Europeans and Asians in North America influences the culture of science there. The motivations and the drives are very, very similar: we all want to do important, interesting work.

The research community is a mix; people come from everywhere. Within TLL, the principal investigators come from North America, from Europe, and from Asia. It is quite diverse.

ARF: How does research funding work in Singapore?

SC: Like other countries, Singapore has a mixture of funding opportunities. There are core-funded research institutes; TLL is one of those. Researchers who are in the university will rely more on grant funding. People who have good ideas, and good projects—and who can write a decent grant—will get the funding they need to do their work.

ARF: Do you still feel connected to your colleagues who are now in very far off time zones?

SC: The world is small! E-mail is tremendously efficient. We have discussions over the Internet, using Skype, for example. There is nothing better than writing a paper with somebody in the U.S. You can work all day on a manuscript and send it to your colleagues in the States. They can work all day on it there, they send it back, and you have it fresh the next morning, ready to go. I don’t feel the slightest bit isolated here.

ARF: What are your goals as director of this laboratory?

SC: Our goal for TLL is to have it be a center of research excellence in this part of the world. My goal is to hire really talented young people, have them excel in their research careers here, and do work that is internationally recognized. When people look to this part of the world, I want TLL to be on their map of places where excellent work is done, and where they might consider wanting to come and do their own work.

ARF: How do you see the role of microRNAs in aging?

SC: There are hundreds of microRNA genes in animal genomes. Collectively, they target a very large fraction of the protein-coding transcriptome. I suspect that for any biological process you chose to name, you would find a microRNA that was involved, in some interesting way, in regulating it.

A very large proportion of microRNAs have a significant fraction of their total expression in the brain. Undoubtedly, there will be roles in aging. Undoubtedly, there will be roles in neurodegeneration. There may be roles in specific diseases. But it is early days in understanding these specific functions of individual microRNAs in these complicated processes.

ARF: How did you get interested in this topic?

SC: I was not planning to work on microRNAs. We were studying genes that are involved in linking growth and patterning, and stumbled onto microRNA genes. From the chance findings of studying a particular microRNA mutant, we were led into looking at the role that this microRNA had in cell death in the brain. The particular target that the microRNA regulated in that instance was a gene—atrophin—which had been identified independently as being involved in human glutamine expansion diseases.

ARF: This would be miR-8?

SC: Yes. A loss-of-function mutant was made in the microRNA miR-8. These animals tended to die before reaching adulthood, and had some minor morphological problems. But when we looked at the surviving animals, we observed behavioral abnormalities that became more severe with age.

ARF: What behavior was getting worse?

SC: Flies, in an enclosed space, will climb upward. That’s called negative geotaxis. If a fly brain is not working well, its perception, its orientation in space, and its coordination can be impaired. The assay for that is dead simple—you measure how long it takes the fly to climb up a tube—but it requires a fairly robust, complex array of functions in the perceptual processing and output parts of the fly’s brain.

Mutant flies started out doing worse than normal flies and got even worse with age. We saw elevated cell death in the brain. It was easy to make a connection between something causing neurons to die and impaired function in the CNS—no magic there.

ARF: How did you link miR-8 to atrophin?

SC: We used a combination of two things. One was computational target prediction. The second was expression profiling to look at changes in target RNA level. We looked at the overlap of those two lists: hundreds of predicted targets, hundreds of RNAs change. The lists had only a small number in common. One of them was atrophin, which, because it had already been linked to human disease, was an obvious gene to pay attention to. We were able to show that these defects result from overexpression of atrophin in the microRNA-deficient cells.

ARF: How did you show that?

SC: If you think that the cause of the defect is that the microRNA is unviable to limit expression of the target, then by overexpressing the target you would expect to reproduce the defect. That is, if there is too much atrophin in those miR-8-deficient cells, then if we put in too much atrophin, do we see the same phenotype? The answer was yes. Now we flip it over: if we reduce how much atrophin those cells can make, we would expect to make it better. And we did.

ARF: How is your research group pursuing the role of microRNAs in neurodegenerative disease?

SC: The miR-8 case got me thinking about the potential that all of the other CNS-expressed microRNAs might have to help us identify protein-coding genes that, when misregulated, could cause problems in the nervous system. Atrophin had been implicated in human disease, so it was pretty evident that that was worth paying attention to. Had we been at this point without knowing that, we could have identified atrophin as a disease gene.

It is that line of reasoning that motivates me to want to look at all the other microRNAs, in the hope that there will be target genes, like atrophin, that we don’t already know about. We are now systematically mutating all of the microRNAs in the Drosophila genome and will analyze the mutants for phenotypes of all sorts

ARF: How many mutants are you aiming for?

SC: There are 128 microRNAs that are conserved in multiple Drosophila species. We already have more than 80 mutants. But it is a long way from having the initial mutants, to verifying them, to cleaning them up so that you can actually do the studies, and so forth.

ARF: What have you discovered so far?

SC: For example, there is a central nervous system-expressed microRNA; the mutants are homozygous viable—but they just don’t move. You can put them on the table, or under the microscope, without anesthetizing them. The normal fly—ffft! it’s gone. If you gently poke the mutants with a paintbrush, they’ll move a little bit, but otherwise they just sit there. If you push them off the edge of the table, they fall to the floor.

It could be a defect in the neuromuscular junction. It could be a defect in the motor output system. It could be a defect in some aspect of central processing. It could be a defect in proprioception: they may not know where their limbs are, so they cannot figure out what to do with them. We have absolutely no idea what is wrong with these guys; it is far too early to say. But it is just one example that illustrates that interesting phenotypes are going to come out of this.

ARF: In your review last year (Bushati and Cohen, 2008), you cited a variety of studies potentially linking miRNAs to neurodegenerative disease or neural health, but ultimately said that it remains uncertain if they play an active role in disease etiology or progression. What would it take to convince you?

SC: It is still early days in this field; a lot of the data are correlational. We need direct evidence that misregulation of a target, or set of targets, by a microRNA, causes the defect. It is just a matter of time until people dig more deeply into functions of individual microRNAs, or identify disease-associated relationships that point to microRNA target involvement. The causal links are coming; it just takes time.

ARF: In that case, could microRNAs or drugs that target them be potential therapeutics for a disease like Alzheimer’s or Parkinson’s?

SC: It would be naïve to say, yes, this holds out great hope for the future of our aging brains, because it is not going to be that simple. It depends on what the problem is. If the problem is due to overexpression of a microRNA, you could imagine therapeutics that target a specific microRNA to reduce its expression.

Finding correlations between particular conditions and patterns of gene expression gives you diagnostic tools as well. The microRNA expression profile of a cell is a molecular signature that reflects something about the state of expression of the proteome. Those molecular signatures are very useful in diagnostics.

ARF: What are the main questions you would like to see answered about the links between miRNAs and neurodegeneration?

SC: The key thing is to understand the normal functions of the microRNAs, in their normal biological context. What does each individual CNS-specific microRNA do? What are its targets? What are the consequences of removing it? Are any of those consequences of removing it plausibly linked to an abnormal condition in the brain? Can we figure out an interesting way to do something about the causes of that condition? There is a whole category of problems there.

ARF: Do you expect to stay in Singapore?

SC: I expect to stay here for the foreseeable future. I like working here. It is a fascinating city. I think there is great opportunity, for young scientists as well as established scientists, here. As long as that mix stays fun and interesting, I will be here.

Amber Dance Interviews Stephen Cohen

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