Saturday, October 28, 2006

Classic papers in quantitative biology

The hot trend in biology education right now is to train undergrads, grad students, and postdocs in not only biology, but also in enough math, computer science, and physics so that they can function successfully in the highly interdisciplinary research fields that are starting play a more important role in current research. Most major research universities are spending serious time and money thinking about how to help physicists and computer scientists become competent in biology, and biologists become competent in the relevant physics and computer science. We have a program for this purpose here at Washington University.

Biologists at Princeton have been experimenting with a variety of educational approaches, and in this month's issue of Nature Reviews Molecular Cell Biology has an essay about one course at Princeton that focuses on what I'd call 'classic papers in quantitative biolgy.' They put their grad students in a room - physicists, biologists, etc., all interested in biological problems, and have them discuss papers that contain not only great biology, but great examples of quantitative work as well.

Anyway, in this essay, there is a list of 12 of the best papers used in the course. If you're interested at all in genomics or what's being called systems biology, I highly recommend checking out this essay and the list of papers. Unfortunately you need some sort of university library access for most of them, but they're worth the effort to get them.

Thursday, October 26, 2006

Comparing Genomes on a Chip

Last week at our lab meeting I had a chance to hear a presentation by Doug Berg, a microbiologist here at Washington University. Berg's work is a great combination of new technology, genomics and evolution, and it happens to also have potential medical relevance. He's studying the evolution of drug resistance in Helicobacter pylori, a ususally benign bacterium that is responsible for stomach ulcers. (Recall that the Nobel Prize in medicine was awarded last year to Barry Marshall and Robin Warren for their discovery of the link between H. pylori and ulcers.)

Berg is basically evolving highly drug resistant bacteria in the lab and using a new genome comparison technology to identify the genes that are changed as the bacterial strains become more resistant. The drug used in these experiments is metronidazole - a substance that itself is not harmful, but inside a bacterial cell it is metabolized into a toxin that causes significant DNA damage.

To create these drug-resistant strains, bacteria are spread on an agar plate - a peteri dish filled with a Jello-like substance that contains nutrients and metronidazole. Most bacteria are killed by the drug, but some individual bacteria survive, if they harbor random mutations that just happen to confer drug resistance. These surviving bacteria continue to divide, and after a few days a colony - a blob of bacteria all descended from the original surviving bacterium - has formed on the plate. You can pick off this colony with a toothpick and save it; you now have a drug resistant strain of bacteria. This process can be repeated - grow up the drug resistant strain, smear it on an agar plate with a higher concentration of the drug, and watch for the next round of colonies to grow from the survivors.

After doing many experiments like this, the Berg lab gets a series of H. pylori strains, each with varying drug resistance. The obvious question then is, how do the genomes of these strains differ? What specific (but randomly occuring) mutations had to take place in order for this level of drug resistance to evolve? Sequencing the genomes of each of these various drug-resistant strains would be the obvious way to find these differences, but doing that kind of sequencing is still prohibitively expensive and computationally intentive. But the Berg lab is using a new technique that rapidly compares genomes on a chip. (The link leads to an abstract of their paper; the full text is subscription only but you can check out more about the technique in a "webinar" found here, at the bottom of the page.). They use microarrays (DNA chips) that cover the entire genome (called a 'tiling array'). By hybridizing DNA from both the nomral and drug resistant strains to the microarray (check out my earlier summary of how a microarray works), they can identify those spots on the array (and thus in the genome) where the two strains differ.

Using this technique, the Berg lab has identified key genes involved in H. pylori resistance to metronidozole. There seem to be some genes that are always changed in these drug resistant strains, and some that can be different. In other words, there seem to be multiple evolutionary paths to drug resistance, but those paths often cross.

This techique of comparing genomes on a chip has a lot of potential, and within a few years, something like this may start showing up in common medical care and hospital labs, replacing the more primitive techniques that are currently used for classifying tumors or drug-resistant pathogens.

NY Times feature on science fraud

The NY Times has a feature article about the first case of scientific fraud with NIH money that resulted in a jail sentence for the scientist involved. A young technician in the lab of Eric Poehlman risked his reputation and future career to expose years of fraudulent research in the Poehlman lab. The grad students and postdocs in the lab were willing to look the other way for years, even though they knew the data that was ending up in the lab's published paper's sometimes came straight out of Poehlman's imagination.

I'm glad Poehlman went to prison. Someone who commits massive fraud like that wrecks trust within the scientific community and with the greater public. Without trust, between colleagues, and between the public and the scientists paid by the public, the whole merit-based funding system is threatened.

Friday, October 20, 2006

Can you learn lab skills with a keyboard and mouse, instead of pipettes and test tubes?

Today's NY Times features an article about a conflict among educators over online lab courses. Universities are debating whether to accept virtual lab courses as a substitute for real Freshman science labs, especially when it comes to AP credit.

Online schools allow students to do virtual pig dissections, virtual DNA gels, and virtual chemistry experiments, and defenders of the vitrual lab courses argue that their students do just as well, if not better on AP exams. If your goal is a high exam score, then I don't question that these virtual labs can help students effectively learn the concepts. But science is more than just concepts, and virtual labs can never take the place of real labs in seriously training scientists.

Learning to be a scientist is much like an apprenticeship - you could never learn to be a master chef, violin maker, or serious gardner if you tried to learn all your skills in an online simulation, even though that simulation might help you learn the concepts involved. You can't learn to be a serious mechanical engineer if you never set foot in a machine shop. Experimental science requires something like a green thumb - a hard to desciribe ability to use your tools with just the right touch. Many experiments are technically challenging, and require dexterity and experience to do them well - that is, reproducibly. (And, going back to the NY Times article, the 'kitchen chemistry' they describe is no substitute. Kitchen chemistry is usually just a bunch of magic tricks; it doesn't come close to teaching kids what real experiments are like.)

Actually, I think this point is generally true of all online learning that tries to be a substitute for a real univerity education. All serious scholarship, whether it involves lab work or not, requires an element of apprenticeship to learn. At most, virtual courses can help students learn the basics, but a complete online university is more like a technical school or certification program, not a real university.

But what about kids in poor schools that can't afford to have serious science labs for their AP courses? In those cases, a good virtual lab is probably better than a poor or nonexistent real lab. Universities have to decide, whether all of their graduates need that kind of lab experience. For students graduating in science or medical fields, the answer is an unequivocal yes. For the others, I don't know the answer. But, in light of the fact that the US is producing fewer and fewer home-grown scientists, a great hands-on science lab can be a great way to get the AP students who took these virtual lab classes interested in a science career.

Sunday, October 15, 2006

Reasons to oppose stem cell research? Debunking the opposition to Missouri's Amendment 2

Around my neighborhood here in St. Louis you can find a lot of lawn signs dealing with "Amendment 2" - a stem cell research and therapies amendment which will be on the ballot next month.

As a general principle, I think the public (the taxpayers funding scientific research in the US) should be involved discussing what kind of science should be done in our society. Mind you, they shouldn't be judging the scientific, technical merit of specific proposals - that's as absurd as asking someone with no engineering training to evaluate the structural integrity of a proposed bridge design.

But some general input is good. I just submitted a funding proposal to the American Cancer Society. On their review panels, they include non-scientists who are interested in cancer research, and my application includes an explanation in non-technical language of what I'm proposing to do and how it is related to cancer. I am judged partly on how well I can explain my work to a general audience, and, more importantly, on whether I can persuade the non-scientist on the panel that my work is important for cancer research. I think that's a great part of the process.

On the other hand, often in the political arena, people's arguments on science issues are based on pure dishonesty and bad faith. The opposition movement to Missouri's amendment 2 falls under this category; they are lying to scare people into voting against the amendment. In their flyers and on their website, they lecture us about ethics, but they are liars and deserve no credibility on this issue.

These are harsh words, but as you'll see below, almost every reason they give to vote against this bill is either deliberately misleading or based on a flat-out lie.

To see that these people are liars, you have to know first what the amendment says. It's never a good idea to take someone else's word on what a ballot initiative says, so you read it yourself right here. And if you're not a resident of Missouri, you should still be interested, because many other states are wrestling with the same issue right now, not to mention our national debate on the subject.

Here's my summary of what I think is a very reasonable initiative (and really, check me by reading the initiative yourself):

The Main Point: stem cell research and therapies permitted under federal law shall be legal in Missouri. Since very little is actually legal or funded under current federal law (and this amendment isn't providing any money), right now this makes little difference. This amendment is really aimed at a time when, say, Congress could override a Bush veto of a stem cell bill like the one that was recently vetoed. This amendment is just assuring that stem cell research and therapies are treated like any other research. You don't generally see certain mainstream research or therapies banned in certain states, while permitted overall at the federal level.

Important restrictions specified in the amendment:

- no cloning a human being - that is, you can't implant a human embryo in a human uterus, if that embryo was not created with human sperm and egg. A cloned embryo is created without sperm (referred to below as Somatic Cell Nuclear Transfer or SCNT).

- no creating a blastocyst purely for research by fertilization - in other words, embryos taken for stem cells would be left over at in vitro fertilization (IVF) clinics, or created through producing a clone by SCNT (if permitted by federal law).

-no taking cells from a blastocyst after more than 14 days of cell division

-no selling eggs or blastocysts

The amendment also includes typical provisions that already apply to all ongoing research using human subjects or samples; these provisions are redundant, because they are already required by current federal laws and regulations:

- donors of eggs/blastocysts must give informed, voluntary consent
- researchers must comply with normal standards of bioethics, must have Institutional Review Board approval (required at all research universities before someone can work with human subjects or tissue samples), and comply with all other regulations that apply to research with human subjects and samples in general.

This proposal basically deals with all non-fundamentalist ethical issues, and I predict that such an approach will be adopted at the federal level within 4 years (and maybe even during the next Congressional session).

A side note on cloning: Beware of people who try to tell you cloning is cloning is cloning - and that there is no difference, it's all ethically the same. Not true! Scientists use the term cloning to mean different things. Cloning a gene is not the same a cloning a human being. Molecular biologists make copies of genes and put them into a variety of different contexts; they call that cloning. Making a human clone involves implanting a cloned embryo into a uterus, and having the woman eventually deliver the baby. This has not been done yet, and it's unethical because you would probably have hundreds of women suffer hundreds of miscarriages before you actually had a live birth. On top of that, that live child will probably have future health problems (because that's what we've observed in cloned animals). But these human clones would not be soulless robots (the Hollywood-inspired fear expressed by a woman I met on the train the other month) - they would be like the natural clones among us today - identical twins, fully autonomous human beings.

Cloning embryos for stem cell research starts out the same as human cloning (pull the DNA from an egg, put in DNA from an adult cell, and get it to divide without fertilization by sperm - SCNT), but you don't implant the embryo in a uterus. People worry that doing this though is the first step towards human cloning, and what's to stop us from going all the way? Well, the fact that cloning an embryo is the easy part, getting it to develop in the uterus is the really hard part. And also, the ethical issues significantly change once you implant such an embryo in the uterus, as I stated before.

Back to amendment 2: the proposed amendment is facing a disinformation campaign by a dishonest opposition. The "Missourians Against Human Cloning" have put out 20 talking points, each of which is either severely misleading or an outright lie. If you've had enough of this issue, you can stop reading the post here, but for all the gory details, keep reading:

- "Reason 1: Amendment 2 would permanently change Missouri's Constitution." Permanently, until voters adopt an amendment to change this one. This claim is misleading, because it ignores that fact that, unlike what happens at the federal level, state constitutional amendments are a common way of doing business in most states, and the amendments are easy to change. Missouri has four proposed amendments on the ballot this Fall, most dealing with mundane issues like property tax exemptions and pensions for state officials convicted of felonies. There is nothing unusual about putting an issue like stem cell research into a state constitutional amendment.

- "Reason 2: Amendment 2 will use our tax dollars for unethical and unproven research. Despite claims to the contrary, if Amendment 2 passes, Missourians will pay for unethical and unproven research indefinitely." Also dishonest - the amendment provides NO MONEY. Any money for such research would come from the NIH budget, which is unlikely to get an extra raise specifically for stem cell research. If you want to talk about our tax dollars going to something unethical (and, of unproven or disproven effectiveness), let's talk about the Bush administration's torture policy.

- "Reason 3: Amendment 2 would create a constitutional right to devalue human life - it would treat human life as a commodity and raw material for unethical human experimentation by the bio-tech industry." Here's an outright lie - the amendment specifically prohibits treating eggs or embryos as commodities - that is, something to be bought and sold. Beyond that, the claim that stem cell research for disease cures in some vague, unarticulated way devalues human life is weak. Notice the scare language that implicitly evokes Nazi-style research - stem cell research is 'unethical human experimentation'!

- "Reason 4 - Amendment 2 is harmful to women... up to 35% of women who submit to ovarian stimulation experience health consequences, and up to 14% of these are severe." Those numbers are false. The procedure is uncomfortable, but it's not really more risky than an appendectomy. It's done routinely, every day, all over this country. My wife went through many rounds of ovarian stimulation drugs, and went through egg retrieval twice. These drugs have been used for a long time, they have a long track record, and they are safe in the vast majority of cases (no riskier than the anesthesia used in a routine appendectomy). It's extremely common, and I would be money that you know someone who has taken ovarian stimulation drugs, even though you may not be aware of it.

- "Reason 5 - Amendment 2 is unnecessary - adult stem cell research has been proven effective for many illnesses... We need to retain the option to direct our tax dollars to this ethical and promising research." That's false - there are actually few illnesses that adult stem cells work for - anything requiring a bone marrow transplant, and that's just about it. We always have the option to study adult stem cells, and labs who are already doing this work are not suddenly going to drop it once there is money for embryonic stem cell research. Furthermore, the adult stem cell researchers I know personally are some of the most passionate advocates for embryonic stem cell research, even though they have no intention of dropping all of their current work to study embryonic stem cells should funding become available.

- "Reason 6: Amendment 2 protects and promotes human cloning. The proponents want you to believe that Somatic Cell Nuclear Transfer (SCNT) is NOT cloning, but the reality is that every textbook and scientific journal defines SCNT as Cloning." Here they dishonestly try to muddy the waters by exploiting the fact that scientists use the term cloning in different ways - see my earlier note on cloning. The ethical issues relevant to making a live human clone are different from those involving cloning embryos (or SCNT) that will never see the inside of a uterus. You have to articulate each of these ethical cases individually. I can think of reasons why it's wrong to make human clones (health problems in the cloned humans, emotional and physical risks to the women who carry these pregnancies with a high risk of failure), but those reasons don't apply to embryos that are never implanted in a uterus. The Amendment 2 opponents are think they can just get away with screaming CLONING!!! instead of articulating a serious argument against embryo cloning. This is a tactic that repeatedly shows up in this debate.

- "Reason 7 Amendment 2 is an elite initiative" - basically, 95% of the promotional money for this amendment came from one wealthy couple. This talking point is not dishonest, it's just stupid. We'll see how elite this amendment really is when we vote on it next month.

- "Reason 8 - Amendment 2 is immoral. It is ethically wrong to destroy human life regardless of its origin or geography." I'm not sure what geography has to do with anything here, but if Amendment 2 is immoral because it 'destroys human life', then so is the process used for in vitro fertilization and many common forms of birth control. Most citizens don't seem to be outraged over birth control or IVF, so I take it that most people don't buy this ethical reasoning.

- "Reason 9 - Amendment 2 will exploit the disadvantaged" because they will be paid to donate their eggs for research. In reality, this won't exploit the 'disadvantaged' any more than any other research involving human subjects - people who volunteer for such studies are paid a small fee in return for their time, effort, and travel expenses. This is how all new drugs and treatments are tested - if amendment 2 is exploitative, so is essentially all research with human subjects. Many such studies involve procedures much more risky than egg donation.

- "Reason 10: Amendment 2 is wrong. To permanently change our constitution to protect unproven and risky scientific experimentation is wrong." We're only halfway to Reason 20, and already they're recycling their previous reasons (see Reasons 1, 2, and 5). I guess this means that there really aren't 20 independent reasons to vote against Amendment 2.

- "Reason 11: Amendment 2 ignores proven research. Ethical stem cell research provides real hope without cloning or destroying human life..." Now they're recycling Reason 5.

- "Reason 12: Amendment 2 is deceptive - it will PROMOTE not ban human cloning." Recycling Reason 6 here. Like I said above, this is a dishonest and misleading attempt to muddy the various meanings of 'cloning'. Amendment 2 bans cloning human beings, but it does not ban the creation of cloned human embryos that will never be implanted in a uterus. Amendment 2 is clear about it's definitions (again, go read the entire thing yourself); the Missourians Against Human Cloning are the ones being deceptive.

- "Reason 13: Amendment 2 will destroy human life." Amendment 2 could save human lives through the treatments it might facilitate. Unless you think destroying blastocysts is equal to murder, Reason 13 is not true.

- "Reason 14: Amendment 2 will NOT give Missourians more access rights to cures or therapies that they and every other American ALREADY enjoy." This is dishonest - that's not the point of the amendment; the point is to make sure that future therapies permitted elsewhere aren't banned in Missouri.

- "Reason 15: Amendment 2 ignores the success of Adult Stem Cell therapies and cures." Wow, we're recycling Reason 5 for a second time. Amendment 2 does not 'ignore' Adult stem cell research - it doesn't ban it and it doesn't take away research dollars (especially since it doesn't provide any research dollars for embryonic stem cell research). And naturally adult stem cells therapies are farther along than embryonic stem cell therapies - adult stem cell research has been going on longer, and that research hasn't been blocked at the federal level.

- "Reason 16: Amendment 2 will NOT insure First Class status to Missouri's biotech industry." This is, amazingly enough, largely true. There is no generic 'First Class status' - different communities excel in different areas of biotech. Missouri can excel in other areas without being #1 in stem cell research.

- "Reason 17: Amendment 2 is NOT in step with the rest of the world." They go on to say that 27 countries have banned human cloning. But again, they are dishonestly exploiting the multiple definitions of cloning. Some of those countries have passed legislation essentially identical to Amendment 2, which does ban cloning full human beings but allows other research. To use those countries as an argument against amendment 2 is extremely dishonest.

- "Reason 18: Amendment 2 will require your hard earned tax dollars." Recycling again - go back and look at Reason 2. They go on to claim that "private money has been directed to more promising research avenues," which is an outright lie - private money is almost the only source of money currently available for stem cell research in this country. It would be helpful if federally funded labs (meaning, most university research labs) could be involved in stem cell research.

- "Reason 19: Amendment 2 has potential to make some people and corporations very wealthy at Missourians' expense. Researchers and bio-tech corporations stand to make a lot of money from patents - even if no cures ever come from human cloning and embryonic stem cell research." In a sense this is true of all government sponsored research - taxpayers fund research at academic labs, which is then developed into usable drugs and therapies by corporations who earn a profit. The whole thing about patent wealth even if there are no cures - this is a little crazy. Honestly, how many people get wealthy from patents on technologies that don't work?

- "Reason 20: Amendment 2 contains no real internal controls or oversight." Another lie. The internal controls and oversight specified in Amendment 2 are exactly the same as those required for any other research involving human subjects. Universities have to certify to funding agencies that they are compliant with these regulations, every time a researcher submits a grant proposal. There is nothing in Amendment 2 that makes stem cell research exempt from all of the regular oversight that is currently in place for studies like vaccine trials, drug tests, experimental surgeries, etc. Reason 20 does not offer an argument specific to stem cell research.

These talking points are largely based on fear-mongering, dishonest blurring of definitions of cloning, and base appeals against 'elites', the wealthy, and the biotech industry. If you live in Missouri, you should be embarrassed to have a 'Vote NO on Amendment 2' lawn sign (at least ones issued by Missourians Against Human Cloning). If you're not in Missouri, I'm sure you'll see nearly identical arguments at some point in your own state, or if not, again at the federal level once the next Congress is in session.

Tuesday, October 03, 2006

Nobel week kicks off with a well-deserved award for RNAi

It's Nobel week, and I'm enough of a nerd to anxiously look forward to this week every year. Yesterday the Nobel committee annouced that the Medicine/Physiology prize is going to Andrew Fire and Craig Mello. Fire is currently at Stanford, and Mello is at U. Mass in Worcester.

It's great to see the prize go to scientists who are under 50, recognizing work that was done less than a decade ago. When the prize recognizes very old work too often, the Nobel loses much of its excitement and risks losing its relevance to current science. The original Nature paper by Fire, Mello, and their colleagues, came out in Feb. 1998. Since that time, RNAi has become huge in the field of biology - both in terms of its role in the cell, as a powerful genetics tool, and as a potential way to do gene therapy.

RNAi, or RNA interference, is when the expression of a single gene is shut off by the presence of foreign RNA molecules that match part of the sequence of that gene. RNAi occurs in plants, animals, and fungi, and most likely evolved as a defense against viruses and other genomic parasites (such as transposons - they're like viruses that spread through the genome and never leave the cell; we all have a lot of these parasites hanging around in our genomes).

What did Fire and Mello do? A look at their work illustrates how major scientific discoveries don't just come out of thin air - they are based on the groundwork usually laid down by several different research groups. Fire and Mello didn't discover RNAi - scientists had known for years that injecting small RNA molecules into organisms like round worms (such as C. elegans - the organism Fire and Mello work with) could inhibit the expression of single genes. Similar phenomena had been described for both plants and bread mold. (Bread mold - N. crassa - is another weird organism that has been used to study many basic cellular processes.)

People hypothesized that it worked because injected 'anti-sense' RNA bound (or 'hybridized') to the cell's messenger RNA (mRNA) and prevented that mRNA from serving as a template for synthesizing new protein:

(Recall that the letters A,C,G, and in RNA, U stand for chemical groups that match up with each other - A pairs with U, C pairs with G.)

But there were problems with this idea - it also worked with 'sense' RNA as well, which would not bind to the mRNA molecule. Also, a small number of injected RNA molecules, not nearly enough to bind up all of the mRNA produced by a partuclar gene, could cause RNAi - suggesting that the injected RNA molecules were reused over and over to break down the mRNA (that is, they funtioned catalytically). This effect could even been seen in the next generation of organisms, that had not been injected with foreign RNA.

Fire, Mello and colleagues came up with the brilliant idea that the phenomenon might be caused by double-stranded RNA molecules, which would have been present as contaminants in any injected sample of RNA. Such double-stranded molecules would not work by simply hybridizing to the mRNA (as shown in the figure above). The process would have to work by some other mechanism, which at the time was completely unknown.

So Fire and Mello deliberately prepared sense, anti-sense, and double-stranded RNAs, injected them into worms, and looked to see which RNA molecules would be most effective at knocking down gene expression.

They picked a clever target gene to knock down - unc-22, a gene coding for a muscle protein. When that muscle protein is absent, the worms uncontrollably twitch. So Fire and Mello injected the different RNA molecules into worm embryos, and looked to see which ones resulted in twitching adult worms, indicating that the unc-22 gene had been shut off. Their results conclusively demonstrated that it was the double-stranded RNA molecules, not the individual sense or anti-sense molecules, that caused RNAi.

This paved the way for later discoveries figuring out excatly how this process works, and for making huge collections of these RNAs that can be used to knock down just about any gene you want in model organisms like flies and worms. Until RNAi, you couldn't shut off genes so easily, except in microorganisms like yeast (which is one reason why yeast is so useful for studying basic molecular biology). Now, you can do those things in multi-cellular organisms, and hardly a week goes by without the report of some new discovery made using RNAi. These double-stranded RNA molecules are even being tested as drugs in humans, to shut off aberrant genes in people with certain diseases. This was certainly a timely and well-deserved Nobel prize.