The Futile Cycle

The crystal structure of a human GPCR (beta 2 adrenergic receptor)

Crystal structures of membrane proteins are really hard to obtain, because they tend to slide and be too “slippery” to get them to pack into an ordered, structured crystal. GPCRs are even harder, because they’ve got so many moving parts that it’s much harder to prevent them from sliding against each other. This is an amazing achievement, but even now you can see that they weren’t able to get everything they wanted; the resolution of the structure is low (over 3 angstroms) and a lot of their protein still didn’t crystalize very well. Still, very impressive.

One of the cool things about biology is that even parasites can have parasites, and those parasites can have parasites…

P2 is a virus that attacks E. coli bacteria (a “bacteriophage”, from the Greek word phagos which means “one who eats”). P2 sometimes lies dormant in E. coli for a while, until for whatever reason it decides to pop out and take over the bacterium to make copies of itself again. When P2 becomes active, its DNA, which was also lying dormant, becomes active and forces the host to make new viruses. The P2 DNA carries the blueprints for the proteins that form its outer coating, which lets it infect other bacteria.

Viruses are really just tiny parasites that taking control of bacteria and force them make new viruses.

P4 is another bacteriophage, but it’s kind of crappy; it doesn’t have genes for its outer coat, so it can’t even make copies of itself! If there is a P2 lying dormant in the same cell, however, P4 will commandeer the other virus’s genes and use its head and tail proteins for itself to make new P4 viruses (instead of P2 viruses). So P4 is really a parasite of P2, which is itself a parasite of bacteria!

Another parasite of a virus is SaPI1, a “pathogenicity island,” which is a piece of DNA found in Staphylococcus aureus, a bacterium that’s responsible for tons of deadly infections these days in schools, hospitals, and prisons. Staph is one of the leading killers of people in the US. But that’s a whole different topic unto itself…

Anyway, pathogenicity islands are pieces of DNA that can cut themselves out of bacterial DNA and reinsert in other parts. One pathogenicity island related to SaPI1 encodes for the protein that causes toxic shock syndrome in humans when they get a staph infection. Normally, pieces of DNA aren’t infectious or anything.

SaPI1 is an interesting island, in that it can actually get out of bacteria and infect other ones! It’s like a virus, but again, it can’t make the virus coat proteins it needs to actually infect other bacteria. But what it can do is take control of a dormant 80-alpha virus! SaPI1 forces 80-alpha to make coat proteins for the SaPI1, which then leaves the current host and goes to infect other cells. So, SaPI1 is a parasite of 80-alpha, which is a parasite of S. aureus, which can infect humans.

In the end, I guess we’re just at the bottom of a whole ‘nother food chain. But it’s a really awesome food chain!

This week I will most likely not be posting, mainly due to my taking my “Molecular Biology of Prokaryotes” exam. It reminds me of the theoretical mathematics class I took as an undergrad; most of my time was spent staring at an empty page, waiting for inspiration to hit (or as Douglas Adams puts it, “until your forehead bleeds”).

As one of the professors here said, “When I read through the midterm, it was not obvious to me how to solve the problems.”

It’s not every day you see a painting swarming with bacteria; or in this case, a painting made with bacteria: (via Wikipedia)

It’s a painting of a beach by Roger Tsien’s lab. They inserted differently “colored” fluorescent proteins into a bunch of bacteria, and then “drew” the picture with toothpicks that had been dragged through the colonies of those bacteria. Since it’s a little hard to see exactly what you’re doing on a semi-transparent gel (which is what the picture was drawn on), this is actually not that easy to draw!

Reminds me of Lite Brite. Anyone else have those as a kid?

This PhD Comic hits too close to home, especially since I’m applying for an NSF fellowship right now. For the planned research summary, I have to supply keywords, background, detailed methodology, anticipated results, broader impacts, and citations and references all in two pages of 12 pt. Times New Roman font, with 1 inch margins.

But since I’m begging for money, I guess I can’t really complain too much. I’ve learned a lot of tricks for beating the page limit, though. Hyphenating long words at the ends of lines is really key; it can save you several lines of text. The most painful thing to include in the proposal is a citation to a book with a really long title, because there’s really no way to get around putting the entire title in the citation. The long title, however, almost always wraps the text to the next line, taking up two whole lines…

Not to generalize from one anecdote, but Doris Lessig, the 2007 Nobel Laureate in Literature, is a rather testy author. Is it just me, or are a lot of famous, “literary” authors annoyed by publicity? Maybe they’re shy? Consider the excerpt:

I asked [Doris Lessig] if she had heard the news and when she said no, I shared it with her.”Oh Christ!” she said in an exasperated tone that I certainly was not expecting. “[Speculation that I might win the prize] has been going on now for 30 years, one can’t get more excited than one gets.”….”Isn’t this a recognition of your life’s work?” I persisted.”Yes, it is. See, you’ve said it all for me,” the feisty and prolific author responded, turning to head indoors….Clearly, she was not going to make it easy. I managed to get her to turn around when she was half-way up the garden path by asking her if prizes meant anything to her, as obviously they were not the reason she wrote books.”Look, I have won all the prizes in Europe, every bloody one. I’m delighted to win them all, okay?”

A lot of science is done in little glass and plastic vials, mainly because there are some things that are just too hard or unethical to do in live organisms. We even have words for the distinction between science done in a test tube (”in vitro”) versus done in a living system (”in vivo”).The gold standard, of course, is “in vivo”. You want data in vivo, because, frankly, a test tube full of chemicals (even chemicals extracted from cells) isn’t a substitute for a real living system.But really, what is “in vivo” depends on your point of view. If you’re a chemist, throwing your favorite molecule on a bunch of cells in a Petri dish is probably “in vivo” for you; after all, there’s living stuff there. But if you’re a doctor, “in vivo” means “in an animal.” Cells in a plate is just “in vitro.”And in reality, whether “in vivo” is a gold standard isn’t that clear cut. Sure, eventually you wan to see whether something happens in an actual living system, but what if you want to test whether two molecules physically interact (we’re not talking regulation here)? Then the “gold standard” would be an “in vitro” assay (i.e. chemicals in a tube), since in a live cell, there’s all sorts of things that can really jiggle up the FRET, colocalization, yeast two-hybrid, ELISA, or whatever other test that biologists and chemists like to use in living systems. There’s too much other “junk” in a living cell to really tell for sure. So you have to do it in a test tube, where you can control all the conditions.Just a though, the next time someone sneers at an “in vitro” experiment. Just smile and tell them “in vitro” is the new gold standard!

I was recently talking to a faculty member in my department who is working on recruiting underrepresented minority students to apply to graduate school. She mentioned that the NIH was really riding graduate schools (such as Harvard, Yale, and Stanford) hard to increase their ethnic diversity, since right now they’re mostly composed of whites and Asians.

At least in our department, the balance between men and women is pretty even, but the ethnic diversity is essentially nil. There are whites, Asians, and Asian-Americans. A few Southeast Asians round out the total. Almost no African-Americans (or Africans) that I’ve seen (maybe a post-doc somewhere?), maybe a few of Hispanic descent. No native Americans that I can recognize (though I often find it difficult to tell).

The Ivory Tower really is a sea of ivory faces.

But that’s apparently the way it is in all biology departments across the nation. The professor told me that across the nation, there were only 500 African-American applicants to graduate schools in biology programs. 500 in the entire nation. Looking at the ETS numbers, nearly 40,000 students taking the GREs in a year said they wanted to enroll in biology or biomedical research graduate school. Let’s say only 25,000 students actually applied to graduate school, to be conservative (that’s less than two thirds). African-Americans would make up only 2% of the student pool (and these are just applicants). There’s something definitely amiss with this situation.

That’s not something that can be fixed by the faculty at one university department or another. The government needs to do something, because this is a collective action problem. The NIH shouldn’t just pressure the graduate schools; they should throw money at the situation, if they really feel there’s a problem (and I think there is). Establish scholarships. I mean, with $10 million, they could establish 100 fellowships that fund at the level of the NSF or DOD fellowships, giving 3 years of a $30,000 stipend. Maybe they could even cut back the war spending 0.01% and use that money to provide 500 fellowships. The situation will be difficult to change for any one university; just putting the pressure on them will make them squabble over the 500 prospective students that are out there.

As everyone probably already knows, the 2007 Nobel Prize in Medicine or Physiology went to Mario Capecchi, Martin J. Evans, and Oliver Smithies for developing a way to create transgenic mice by targeting genes. Smithies and Capecchi developed the methodology for creating cells which had specific genes knocked out, while Evans developed a lot of the methodology for using these techniques in stem cells to create live mice that would carry these induced mutations. The knock-out mouse is now one of the most versatile models we have for testing questions about genes and diseases. A good explanation of the process that they developed is explained here.

I know a lot of people just skim the press release, but there are some really interesting nuggets in the “Advanced Information” section of the announcement. For example, Evans first tried to use cancer cells to make mice, but (as you would expect), the cells were just too sick to actually make a good organism. So he actually went through and found the cells the we now use as “Embryonic Stem Cells,” which he used to create mice that were a mosaic of cells of two different mice.

Another thing was that Capecchi and Evans both wrote grants proposing this research (on gene targeting) to the NIH and the UK Medical Research Council (respectively), but both were rejected, because the reviewers thought it would be too hard and unlikely to succeed!

Finally, though this isn’t mentioned on the Nobel website, Capecchi had a really hard life as a child. His mother was arrested and put away in a concentration camp during World War II (for being an anti-Fascist bohemian), and he was left alone for four years as a street urchin. Fortunately, he was later found (very sick and malnutritioned) by his mother (who survived). He moved to the US, went to school, then college, worked under Jim Watson at Harvard, and then went on to have a massively successful career (though, as I mentioned above, not without a few hiccups along that path as well). It’s an amazing life story, and the Nobel Prize surely can’t go to a better person.

It still amazes me how, back in the “old days”, discoveries of base-substitutions, frame-shift mutations, deletions, the genetic code, nonsense mutations, transposons, and gene mapping were done all without the availability of DNA sequencing, which started to come into major use in the mid-1970s. For example, the discovery that transposons (or “jumping genes”) were surrounded by DNA sequences of inverted repeats was done not by DNA sequencing, but by electron microscopy. Yes, electron microscopy. Researchers concluded after seeing that the transposons could form cruciform stem-loop shapes that they had such inverted repeats.

How cave men might have looked for inverted repeats

This is an amusing paragraph:

“[One researcher] found [transposon-like insertions] which do not have detectable inverted repitition….These insertions may represent a different type of [transposible] element, or may be cases in which the inverted repetition is too short or too unstable to be detectable by standard visualization methods which depend on intramolecular annealing. [i.e. electron microscopy]” (from Kleckner et al. (1975) J. Mol. Biol., 97, 561-575.)

Seriously, this paper sounds like something from the dark ages. Nowadays, any reasonably skilled undergraduate would be able to just sequence the darn thing, showing without a doubt whether the inverted repeats are there or not.

Just reading these old papers makes me sympathize with the researchers in the 1990s who wanted to sequence the hell out of everything they could find (and they’re still out there). I mean, when compared to technology like this, sequencing must have seemed like the be-all-end-all of molecular biology!