Huttenhower Lab, 2017-Present

Stay tuned - I'm still sorting out what my project will be.

Dutton Lab, 2014-2017

Publications:
Kevin S BonhamBenjamin E WolfeRachel J Dutton. "Extensive Horizontal Gene Transfer in Cheese-Associated Bacteria" eLife, (2017) http://dx.doi.org/10.1101/079137
Work by Undergraduates:
Invasion of the cheese rind community byListeria monocytogenes

Microbes from cheese rinds can be readily isolated and cultured in the lab, and then used in vitro to study microbial communities

Microbes from cheese rinds can be readily isolated and cultured in the lab, and then used in vitro to study microbial communities

The Dutton Lab studies the microbes that grow on cheese, which is super fun to tell people about at parties. But the reason to study cheese goes far beyond an excuse to have delicious treats at lab meetings - we're studying communities of microbes. If you've been paying any attention, you've probably heard about the human "microbiome," the consortium of tiny critters that covers every body surface, packs our intestines, and seems to play important roles in everything from diabetes to depression. That's one microbial community that's clearly important, but complex multispecies microbial communities are everywhere (and everywhere important) from the soil under our crops, to the oceans producing our oxygen, to trash heaps and bioreactors. The trouble is, they're hard to study.

That's where the cheese comes in. The outer surfaces of aged cheeses (called the rind) are complex multispecies microbial communities. But crucially, this microbial community is much easier to study than those of an animal gut or soil. It's a model system - just like E. coli has been a model for molecular biology, or S. cerevisiae is a model to study cell biology, the microbial communities on cheese are tractable systems to study principals of microbial ecology.

My project was looking for horizontal gene transfer (HGT) in our cheese microbes. Bacteria are asexual - when they reproduce, they split into two more-or-less identical copies. In principle, this cuts off mixing of genomes - all genes are "vertically" inherited, in other words, passed directly from parent to offspring. In practice though, a lot of mixing really does occur. Whether because of bacteria viruses (phages) accidentally packaging one victim's genes and pulling them along to another, or through conjugation (this is the closest thing to bacteria sex), or some other mechanism, bacteria are constantly sharing their genes with one another.

The HGT shared between the ~150 bacterial genomes we looked at.

The HGT shared between the ~150 bacterial genomes we looked at.

Precisely why or how or even how frequently this happens is not entirely clear. One reason it's unclear is that it's difficult to study dynamic processes like this in "the wild" (the fancy term is in situ). What you'd really like is a way to bring the community into the lab to manipulate it in vitro, and that's what the cheese ecosystem allows us to do!

I was also interested in HGT as a window into what's important in this microbial community. See, as tractable as the cheese microbial communities are, we're still missing a lot of traditional tools to study them. We might be able to convincingly show that there's an interaction between two species, but showing what the interaction is is a bit trickier. So I took a genomic approach - using computational analysis to sift through all the genes of our bacteria to see if I could identify patterns that tell me something about what the bacteria think are important. 

One way to do this would be to look for genes that are enriched in cheese genomes compared to non-cheese genomes of related species, but doing this with anything approaching statistical rigor would require a lot more genomes than we had access to. Evolution can take a long time, and small differences that have an impact may be tough to pull out from the noise. Instead, I looked at the HGT. The rationale here is that, while the core genomes of these microbes are going to look mostly like their close non-cheese relatives, there may be genes with super important functions that are being swapped around at a high frequency. It's much easier to find a gene that's brand new than it is to find something that's just a bit different. 

These are all the groups we saw, plotted against the average number of horizontally transferred genes per species that has the group.

These are all the groups we saw, plotted against the average number of horizontally transferred genes per species that has the group.

So I wrote a software package to scan through the genomes of our microbes and pull out genes that are likely to be horizontally transferred. What this means in practice is that I compared each genome to a bunch of others, and if the software sees a gene that's super close between two species - much closer than you would expect based on their common ancestry - it gets flagged. To understand the rationale, consider humans and parakeets. We share a common ancestor with birds about 320 million years ago, but still share a huge number of genes that have been passed down from that ancestor. Of course, those genes have been accumulating mutations along the way, so though we can see that parakeet hemoglobin is related to human hemoglobin, they're pretty dissimilar at the level of A, C, G and T. If we found a gene that was 99% identical in humans and in a bird, we'd probably have to conclude that it was the result of horizontal gene transfer (though beyond intentional genetic manipulation, I don't think there's any known way for something like this to happen). That's what I looked for in our cheese microbes - genes that are 99% similar in species whose genes should be much more distantly related.

And I found a bunch! A lot of them seem to be involved in acquiring nutrients, particularly iron. Our paper detailing this work was published in eLife in June of 2017, and can be found here.

A figure from Bonham et. al. Cell 2014 showing the localization of TIRAP (green) compared with something localizing only to the plasma membrane (red, overlap = yellow)

A figure from Bonham et. al. Cell 2014 showing the localization of TIRAP (green) compared with something localizing only to the plasma membrane (red, overlap = yellow)

I earned my Ph.D in the Kagan Lab, where the focus is innate immunity. There are a ton of projects in the lab, but all of them are (or at least were when I was there) organized around the theme of interactions between cell biology and the detection of infectious microbes. That is, how does the structure of a cell and its spacial context influence our interactions with pathogens?

In order to respond to potentially dangerous microbes, our immune system needs to know that they're there. Cells don't have eyes, but use proteins called "Pattern Recognition Receptors" (PRRs) to detect and bind to biochemical queues called "Pathogen Associated Molecular Patterns" (PAMPs). Triggering a PRR usually mobilizes an immune response, causing inflammation, recruitment of other immune cells, and hopefully a generally bad time for whatever is infecting us. But context is everything - a bacterium inside the lumen of the gut usually isn't a threat, and we want to respond differently to a virus in the brain than to a parasite in the blood. The cellular context in which a microbe is detected has a great deal of influence on how the cell responds to the threat.

My project focused on a protein called TIRAP, which is involved in transmitting signals from a class of PRRs called Toll-like Receptors (TLRs). Once the TLR binds to its ligand, the signal has to be propagated into the cell in order to trigger a change in response. TIRAP acts like a bridge between the receptor and the signaling enzymes inside the cell - specifically, it sits positioned at the cell membrane specifically where recognition is likely to occur, and pulls the other signaling machinery to that location. 

Some TLRs hang out at the cell surface, and some hang out in endosomes, but before my project, TIRAP was thought to only participate in signals from the plasma membrane. There was good reason to believe this - if you delete the gene for TIRAP from cells, they can still signal from endosomes, but not from the cell surface. Case closed right? Wrong. It turns out that the molecules originally used to study the phenomenon had a unique quality that masked the requirement for TIRAP. When I used natural ligands of endosomal TLRs (viruses), it turned out TIRAP was required.

Part of the belief in TIRAP's exclusivity was also based on the fact that its localization domain bound PIP2, a lipid only found on the outer membrane of cells. I also showed that, contrary to previous belief, TIRAP could bind to multiple different types of membrane lipids, including some found on endosomes, and this feature allowed it to be in multiple places at once. 

Mowen Lab, 2006-2008

Publications:

Effects of a novel arginine methyltransferase inhibitor on T-helper cell cytokine production.” FEBS J. 2010 
NIP45 controls the magnitude of the type 2 T helper cell response.Proc Natl Acad Sci U S A. 2010

The Mowen Lab at The Scripps Research Institute studies the role of arginine methylation in the immune system. Arginine methylation is a "post translational modification" (PTM) of proteins - that is, it's a mechanism of altering the function of a protein after it's been made. Like its better studied cousin phosphorylation, arginine methylation can activate or deactivate proteins and have profound effects on cellular signaling pathways.

At the time I joined the lab, it was clear that this PTM was important in the development of T helper cells, and perhaps many other immune functions, but there weren't great tools to study it. My project was to characterize a set of small molecule chemical inhibitors of the enzymes (PRMTs) that carry out arginine methylation in the hopes that they could be used to more thoroughly study the role of arginine methylation in the immune system.

Aroian Lab, 2004-2006

Publications: 

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The Aroian Lab studies the effects of different drugs on parasitic nematodes. When I was in the lab, the focus was on toxins produced by Bacillus thuringiensis, which have been made famous as pesticides sprayed on organic crops or engineered into genetically engineered crops to fight insect pests. But it turns out there are also variants of these Bt toxins that target roundworms, and the Aroian Lab is interested in the potential use of these toxins to treat human and agricultural pathogens.

While I was in the lab, one of these toxins (Cry5b) was known, but my project was to screen a panel of natural isolates of Bt from South America to determine if any of them had nematocidal activity. I'm not sure if anything came of my efforts in the lab, but I learned a great deal, and my experiences in the Aroian lab introduced me to the joys as well as the difficulties of doing academic science.