A visiting colleague participated in Monday's lab meeting, and his ideas got me thinking seriously about doing an experiment that I've long claimed wouldn't really prove anything.
The question is whether DNA uptake can provide bacteria with enough nutrients to make a difference to their growth rates. I've always argued that getting some nutrients from the DNA is inevitable, so so demonstrating that this increases growth in a lab culture would not be good evidence that selection for these nutrients is why cells take up DNA. But maybe I was wrong.
In the past we have tried to show that H. influenzae can use nucleotides from DNA for growth, but the experiments have had lots of problems, I think in part because H. influenzae is quite fastidious in its nutrient requirements, and in part because we didn't devote a full-press of our time and brain-power to it.
Our colleague Steve Finkel has shown that E. coli can use nutrients probably acquired by DNA uptake for growth, but the effects are modest. This is probably because the cells are only taking up very small amounts of DNA because their competence genes are not induced under normal culture conditions.
Rather than now trying again with H. influenzae, I'm considering trying to demonstrate nutritional benefits of DNA uptake using Bacillus subtilis. B. subtilis is a soil bacterium, very easy to grow and not at all fastidious. I've worked with it before and one of the post-docs did her PhD work on it. B. subtilis takes up lots of DNA under lab conditions, though not during exponential growth. Competence is induced under conditions referred to as 'post-exponential growth', meaning after cell growth has stalled because nutrients are depleted.
Keeping cells competent over a long period might be complicated, because B. subtilis competence and sporulation are induced simultaneously (though apparently in different sub-populations of the culture). I'd first have to read up on the latest work on the regulation of DNA uptake, and try to find the best conditions for observing a growth effect. But if I can show that B. subtilis cultures grow better when they can take up DNA, I think many other biologists would find the result much more compelling than I do. This would be good, because they are not at all convinced by the results that I think are compelling.
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3 weeks ago in Genomics, Medicine, and Pseudoscience
I'm not sure how helpful this will be, but when I saw this post I thought "flux balance analysis".
ReplyDeleteUsing the H.influenzae metabolic reconstruction, it would be possible to simulate it's growth rate on a variety of media. For example theoretically increase the amount of nucleotide influx into the cytosol and see by how the model predicts a growth rate increase.
I appreciate this might sound a bit far fetched, but it was an idea I had when I read your post.
Steve's group at USC has done really nice work on long-lived cultures. He has a lot of great ideas. I think that you should definitely do the experiment. DNA is a perfectly viable c,n, energy source, and it seems likely that many bugs (B. subtilis being a great example) can use it that way. An interesting thought occurs- a few recent studies (from the Holden lab at UCSB, and at least one other) have shown that DNA is a significant component of at least some biofilm matrices. Therefore, it seems plausible that DNA could be an environmentally relevant nutrient source. Was one of those studies in H. influenzae? I thought it was (I recall a really cool J.Bact cover, with DAPI stained thick cords of DNA in a biofilm matrix). Anyway, I love that you put your research out here in real time, and wish you luck with it!
ReplyDelete"Using the H. influenzae metabolic reconstruction..."? I didn't even know this existed! (I am so out of touch...) I've found the 1999 H. influenzae reconstruction paper and will now start going through it and the papers that cite it.
ReplyDeleteWe've known for about 15 years that there's tons of DNA in many (most?) bacterial environments. There's about 300 ug/ml in healthy resiratory mucus (where H. influenzae lives) and more than 1 mg/ml in people with respiratory diseases. The hard part is getting people to consider that nutrients from DNA might be more valuable to bacteria than rare and random changes in genotype.