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Tag: genetics

Fat Flora

intestines-microflora

November’s paper was published in Nature in 2006, and covers a topic I’ve become increasingly interested in: the impact of the bacteria that have colonized our bodies  on our health (something I’ve blogged about here and here).

The idea that our bodies are, in some ways, more bacteria than human (there are 10x more gut bacteria – or flora — than human cells on our bodies) and that those bacteria can play a key role on our health is not only mind-blowing, it opens up another potential area for medical/life sciences research and future medicines/treatments.

In the paper, a genetics team from Washington University in St. Louis explored a very basic question: are the gut bacteria from obese individuals different from those from non-obese individuals? To study the question, they performed two types of analyses on a set of mice with a genetic defect leading to an inability of the mice to “feel full” (and hence likely to become obese) and genetically similar mice lacking that defect (the s0-called “wild type” control).

The first was a series of genetic experiments comparing the bacteria found within the gut of obese mice with those from the gut of “wild-type” mice (this sort of comparison is something the field calls metagenomics). In doing so, the researchers noticed a number of key differences in the “genetic fingerprint” of the two sets of gut bacteria, especially in the genes involved in metabolism.

imageBut, what did that mean to the overall health of the animal? To answer that question, the researchers did a number of experiments, two of which I will talk about below. First, they did a very simple chemical analysis (see figure 3b to the left) comparing the “leftover energy” in the waste (aka poop) of the obese mice to the waste of wild-type mice (and, yes, all of this was controlled for the amount of waste/poop). Lo and behold, the obese mice (the white bar) seemed to have gut bacteria which were significantly better at pulling calories out of the food, leaving less “leftover energy”.

imageWhile an interesting result, especially when thinking about some of the causes and effects of obesity, a skeptic might look at that data and say that its inconclusive about the role of gut bacteria in obesity – after all, obese mice could have all sorts of other changes which make them more efficient at pulling energy out of food. To address that, the researchers did a very elegant experiment involving fecal transplant: that’s right, colonize one mouse with the bacteria from another mouse (by transferring poop). The figure to the right (figure 3c) shows the results of the experiment. After two weeks, despite starting out at about the same weight and eating similar amounts of the same food, wild type mice that received bacteria from other wild type mice showed an increase in body fat of about 27%, whereas the wild type mice that received bacteria from the obese mice showed an increase of about 47%! Clearly, gut bacteria in obese mice are playing a key role in calorie uptake!

In terms of areas of improvement, my main complaint about this study is just that it doesn’t go far enough. The paper never gets too deep on what exactly were the bacteria in each sample and we didn’t really get a sense of the real variation: how much do bacteria vary from mouse to mouse? Is it the completely different bacteria? Is it the same bacteria but different numbers? Is it the same bacteria but they’re each functioning differently? Do two obese mice have the same bacteria? What about a mouse that isn’t quite obese but not quite wild-type either? Furthermore, the paper doesn’t show us what happens if an obese mouse has its bacteria replaced with the bacteria from a wild-type mouse. These are all interesting questions that would really help researchers and doctors understand what is happening.

But, despite all of that, this was a very interesting finding and has major implications for doctors and researchers in thinking about how our complicated flora impact and are impacted by our health.

(Image credit) (Figure 3 from the paper)

Paper: Turnbaugh et al., “An obesity-associated gut microbiome with increased capacity for energy harvest.” Nature (444). 21/28 Dec 2006. doi:10.1038/nature05414

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Atlantic Cod Are Not Your Average Fish

Another month, another paper, and like with last month’s, I picked another genetics paper, this time covering an interesting quirk of immunology.

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This month’s paper from Nature talks about a species of fish that has made it to the dinner plates of many: the Atlantic Cod (Gadus morhua). The researchers applied shotgun sequencing techniques to look at the DNA of the Atlantic Cod. What they found about the Atlantic Cod’s immune system was very puzzling: animals with vertebra (so that includes fish, birds, reptiles, mammals, including humans!) tend to rely on proteins called Major Histocompatibility Complex (MHC) to trigger our adaptive immune systems. There tend to be two kinds of MHC proteins, conveniently called MHC I and MHC II:

    • MHC I is found on almost every cell in the body – they act like a snapshot X-ray of sorts for your cells, revealing what’s going on inside. If a cell has been infected by an intracellular pathogen like a virus, the MHC I complexes on the cell will reveal abnormal proteins (an abnormal snapshot X-ray), triggering an immune response to destroy the cell.
    • MHC II is found only on special cells called antigen-presenting cells. These cells are like advance scouts for your immune system – they roam your body searching for signs of infection. When they find it, they reveal these telltale abnormal proteins to the immune system, triggering an immune response to clear the infection.

The genome of the Atlantic cod, however, seemed to be completely lacking in genes for MHC II! In fact, when the researchers used computational methods to see how the Atlantic cod’s genome aligned with another fish species, the Stickleback (Gasterosteus aculeatus), it looked as if someone had simply cut the MHCII genes (highlighted in yellow) out! (see Supplemental Figure 17 below)

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Yet, despite not having MHC II, Atlantic cod do not appear to suffer any serious susceptibility to disease. How could this be if they’re lacking one entire arm of their disease detection?One possible answer: they seemed to have compensated for their lack of MHC II by beefing up on MHC I! By looking at the RNA (the “working copy” of the DNA that is edited and used to create proteins) from Atlantic cod, the researchers were able to see a diverse range of MHC I complexes, which you can see in how wide the “family tree” of MHCs in Atlantic cod is relative to other species (see figure 3B, below).

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Of course, that’s just a working theory – the researchers also found evidence of other adaptations on the part of Atlantic cod. The key question the authors don’t answer, presumably because they are fish genetics guys rather than fish immunologists, is how these adaptations work? Is it really an increase in MHC I diversity that helps the Atlantic cod compensate for the lack of MHC II? That sort of functional analysis rather than a purely genetic one would be very interesting to see.

The paper is definitely a testament to the interesting sorts of questions and investigations that genetic analysis can reveal and give a nice tantalizing clue to how alternative immune systems might work.

(Image credit – Atlantic Cod) (All figures from paper)

Paper: Star et al, “The Genome Sequence of Atlantic Cod Reveals a Unique Immune System.” Nature (Aug 2011). doi:10.1038/nature10342

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The Sickle Cell Salve

I might have been crazy late with April, but for May, my on-timeliness when it comes to the paper a month posts is returning with a vengeance.

This month’s paper from Cell (a journal I usually avoid because their papers are ridiculously long :-)) dives beneath the surface of one of the classic examples of genetics used in almost every intro-to-genetics seminar/class/textbook. As you probably know, living things typically receive two sets of genes: one from the mother and one from the father. If those two sets of genes result in the same protein, then the organism is said to be homozygous for that particular trait. Otherwise, the proper term is heterozygous. In classical genetics (i.e. what was painstakingly discovered by Gregor Mendel, the “father of genetics”), being heterozygous, to a casual observer, was usually something that could only be seen after multiple generations (or with a DNA test). This is because even though the individual has two different versions of the same gene, one of them is “dominant”, expressing itself more loudly than the other.

en93587For the mutation which causes the disease sickle cell anemia (see image to the right as to why the disease is called “sickle cell”), however, the truth was a little different. While heterozygous individuals did not suffer from the problems associated with sickle cell anemia, unlike individuals homozygous for the “normal” gene, they showed a remarkable advantage when it came to surviving infection with malaria. It is one reason scientists feel that sickle cell anemia continues to be endemic in parts of the world where malaria is still a major issue.

But, how the sickle cell disease mutation did this in heterozygotes was not well-understood. The authors for this month’s paper tried to probe one possible explanation for this using mice as an experimental system. The interesting thing that they found was that mice that were heterozygous for the sickle cell trait (HbSAD), despite having better survival against malaria than those which were homozygous for the “normal” gene (HbWT) (see the Kaplan-Meier survival curve in Figure 1A below, showing the proportion of surviving mice over time), did not have a significantly different amount of infected red blood cells (see Figure 1F below).

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So if the sickle cell gene wasn’t reducing the number of infected cells, what was causing the improvement in survival? The researchers knew that red blood cells which are heterozygous for the sickle cell trait will often “leak” an iron-containing chemical called heme into the blood stream. Because heme just floating around is toxic, the body responds to this with an enzyme called heme oxygenase-1 (HO-1) which turns toxic heme into the less toxic biliverdin and carbon monoxide (CO). The researchers considered whether or not HO-1 was responsible for the improved ability of the mice to avoid cerebral malaria. In a creative experiment, they were able to show that the sickle cell mice needed HO-1 to get their better survival – mice which were genetically engineered to be missing one copy of HO-1 (Hmox1+/-), even if they were heterozygous for the sickle cell disease, did not survive particularly well when infected (see Figure 2B below, left for the survival data).

In fact, they were even able to show that if you took mice which did not have any sickle cell trait gene (the HbWT group), and replaced their blood system using irradiation and a bone marrow transplant from a heterozygous sickle cell mouse (HbSAD), you only improve survival if the cells come from a mouse with its HO-1 genes intact (Hmox1+/+) (see Figure 4A below, right).

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So, we know HO-1 is somehow the source of the heterozygous mice’s magical ability to survive malaria. But, how? As I stated earlier, the researchers knew HO-1 produced carbon monoxide (CO) and, they were able to show that heterozygous mice with a defective HO-1 response were able to survive when given carbon monoxide (see Figure 6E below). Interestingly, exposure to carbon monoxide reduces the amount of heme floating around in the bloodstream, something which gets kicked into overdrive when malaria starts killing red blood cells left and right (see Figure 6G below), something the researchers validated when they were able to neutralize the protecting power of carbon monoxide by adding more heme back into the mouse (see Figure 6H)

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So, overall, what did I think? First the positive: these are extremely clearly designed and well-controlled experiments. I could only show a fraction of the figures in the paper, but rest assured, they were very methodical about creating positive and negative controls for all their figures and experiments which is fantastic. In particular, the use of bone marrow transplantation and genetically engineered mice to prove that HO-1 plays a key role in improving survival were creative and well-done.

What leaves me unsettled with the paper is the conclusion. The problem is that the trigger for HO-1, what the authors have shown is the reason mice which are heterozygous for sickle cell anemia survive malaria better, is heme, which happens to also be what the authors say is the cause for many of the survival complications. It’s like claiming that the best way to cure a patient of poison (heme) is to give the patient more poison (heme) because the poison somehow triggers the antidote (HO-1).

In my mind, there are two possible ways to explain the results. The first is that the authors are right and the reason for this is around the levels and timing of heme in the bloodstream. Maybe the amount of heme that the sickle cell heterozygotes have is not high enough to cause some of the malaria complications, but high enough so that HO-1 is always around. That way, if a malaria infection does happen, the HO-1 stays around and keeps the final level of heme just low enough so that problems don’t happen. The second explanation is that the authors are wrong and that the carbon monoxide that HO-1 is producing is not reducing the amount of heme directly, but indirectly by reducing the ability of the malaria parasites to kill red blood cells (the source of the extra heme). In this case, sickle cell heterozygotes have chronically higher levels of HO-1.

Both are testable hypotheses – the first can be tested by playing around with different levels of heme/HO-1 and observing how the amount of free-floating heme changes over time when mice are infected with malaria. The second can be tested by observing test tubes full of red blood cells and malaria parasites under different amounts of carbon monoxide.

In any event, I hope to see further studies in this area, especially ones which lead to more effective treatments for the many millions who are affected by malaria.

(Image Credit – Sickle Cell) (Figures from paper)

Paper: Ferreira et al., “Sickle Hemoglobin Confers Tolerance to Plasmodium Infection.” Cell 145 (Apr 2011) – doi: 10.1016/j.cell.2011.03.049

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Do you have the guts for nori?

The paper I will talk about this month is from April of this year and highlights the diversity of our “gut flora” (a pleasant way to describe the many bacteria which live in our digestive tract and help us digest the food we eat). Specifically, this paper highlights how a particular bacteria in the digestive tracts of some Japanese individuals has picked up a unique ability to digest certain certain sugars which are common in marine plants (e.g., Porphyra, the seaweed used to make sushi) but not in terrestrial plants.

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Interestingly, the researchers weren’t originally focused on how gut flora at all, but in understanding how marine bacteria digested marine plants. They started by studying a particular marine bacteria, Z. galactanivorans which was known for its ability to digest certain types of algae. Scanning the genome of Z., the researchers were able to identify a few genes which were similar enough to known sugar-digesting enzymes but didn’t seem to have the ability to act on the “usual plant sugars”.

Two of the identified genes, which they called PorA and PorB, were found to be very selective in the type of plant sugar they digested. In the chart below (from Figure 1), 3 different plants are characterized along a spectrum showing if they have more LA (4-linked 3,6-anhydro-a-L-galactopyranose) chemical groups (red) or L6S (4-linked a-L-galactopyranose-6-sulphate) groups (yellow). Panel b on the right shows the H1-NMR spectrum associated with these different sugar mixes and is a chemical technique to verify what sort of sugar groups are present.

These mixes were subjected to PorA and PorB as well as AgaA (a sugar-digesting enzyme which works mainly on LA-type sugars like agarose). The bar charts in the middle show how active the respective enzymes were (as indicated by the amount of digested sugar that came out):

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As you can see, PorA and PorB are only effective on L6S-type sugar groups, and not LA-type sugar groups. The researchers wondered if they had discovered the key class of enzyme responsible for allowing marine life to digest marine plant sugars and scanned other genomes for other enzymes similar to PorA and PorB. What they found was very interesting (see below, from Figure 3):

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What you see above is an evolutionary family tree for PorA/PorB-like genes. The red and blue boxes represent PorA/PorB-like genes which target “usual plant sugars”, but the yellow show the enzymes which specifically target the sugars found in nori (Porphyra, hence the enzymes are called porhyranases). All the enzymes marked with solid diamonds are actually found in Z. galactanivorans (and were henced dubbed PorC, PorD, and PorE – clearly not the most imaginative naming convention). The other identified genes, however, all belonged to marine bacteria… with the notable exception of Bateroides plebeius, marked with a open circle. And Bacteroides plebeius (at least to the knowledge of the researchers at the time of this publication) has only been found in the guts of certain Japanese people!

The researchers scanned the Bacteroides plebeius genome and found that the bacteria actually had a sizable chunk of genetic material which were a much better match for marine bacteria than other similar Bacteroides strains. The researchers concluded that the best explanation for this is that the Bacteroides plebeius picked up its unique ability to digest marine plants not on its own, but from marine bacteria (in a process called Horizontal Gene Transfer or HGT), most probably from bacteria that were present on dietary seaweed. Or, to put it more simply: your gut bacteria have the ability to “steal” genes/abilities from bacteria on the food we eat!

Cool! While this is a conclusion which we can probably never truly *prove* (its an informed hypothesis based on genetic evidence), this finding does make you wonder if a similar genetic screening process could identify if our gut flora have picked up any other genes from “dietary bacteria.”

(Image credit – Nori rolls) (Figures from paper)

Paper: Hehemann et al, “Transfer of carbohydrate-active enzymes from marine bacteria to Japanese gut microbiota.” Nature 464: 908-912 (Apr 2010) – doi:10.1038/nature08937

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Crazy Y Chromosomes

A few weeks ago, I set myself some 2010 goals. One of which was to make sure that I fit in reading at least one paper every month. What I didn’t say though, was that I would try to do a quick blog post on each (to help keep me honest).

I forgot exactly how I found this paper, but the subject immediately caught my eye:

Chimpanzee and human Y chromosomes are remarkably divergent in structure and gene content

What? I had always been taught that the genomic differences between chimpanzees and humans were extraordinarily small, and since I’m male, I was also drawn to the fact that it would talk about a chromosome that was very near and dear to me.

What I read was pretty amazing. Despite the fact that we are very genetically similar to chimpanzees the Y chromosomes of our two species are actually very different. The best depiction of this that I can point to is from Figure 2 of the paper (below). The two charts below are dot plots which show where the human and chimpanzee chromosomes “line up”, so to speak. The right-hand chart shows how closely related the human chromosome 21 is to the analogous chimpanzee chromosome. You can see this from the nearly perfect diagonal line, showing that the two chromosomes are pretty close to identical as you move from one end of the chromosome to the other. The left-hand chart shows a similar comparison of a human Y chromosome and a chipmanzee Y chromosome. Notice the difference?

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While it was fascinating to read what specifically was different (i.e. repeats, ectopic homologous recombination, introduction of nonsense mutations and open reading frames), what I found most interesting was the speculation as to why the Y chromosomes of two very similar species would diverge so much in such a narrow period of time. The research group’s hypothesis is that the Y chromosome holds a great deal of influence over sperm production, and because a Y chromosome will never have a “partner” chromosome the way that every other non-sex-determining chromosome does, changes in the Y chromosome are likely to have very significant changes in sperm characteristics. Because female chimpanzees oftentimes mate with multiple males, there is strong sperm competition and hence strong selective pressure for chimpanzees to have rapid evolution in the Y chromosome.

Of course, this is all just a guess. One way to test it would be to compare the human Y chromosome sequence with further primate species and see if primates where sperm competition is less intense have more similar chromosomes as humans. Another would be to see if any of the genetic changes resulted in clear sperm/testes genetic or transcriptional control differences.

But, all in all, a very cool start to what I’m hoping will be a fun 12 months!

(Figure 2 from paper)

Paper: Hughes, Jennifer F. et al. “Chimpanzee and human Y chromosomes are remarkably divergent in structure and gene content.” Nature 463, 536-539 (28 January 2010) – doi:10.1038/nature08700

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