eLife Episode 38: Boosting your Intellect
This month in the eLife podcast, how yeast makes an important drug from a plant root, why worms want to kill off males, and are we gender neutral when we pick people to referee papers?
In this episode
00:34 - How bacteria build biofilms
How bacteria build biofilms
with Jeff Boyd and Ameya Mashruwala, Rutgers University
Staphylococcus aureus is a very common cause of infection in humans. In its drug-resistant form - MRSA - it can be lethal. And one of the reasons that Staph. is such a challenge is because once it gains a toehold inside the body it can be very hard to get rid of. For instance, if your hip replacement gets infected, often the only remedy is another trip to operating theatre to remove the entire prosthesis. This happens because bugs like Staph. can assemble protective barriers around themselves called biofilms, which shield the bugs inside from the immune system and antibiotic agents. Now two scientists at Rutgers University, Jeff Boyd and Ameya Mashruwala, have discovered the trigger that makes Staph. aureus begin building its biofilm. Chris Smith spoke to Jeff Boyd...
Jeff - If you’ve ever walked in a stream and you slipped on a real slippery rock, a lot of that slippery surface is actually created by microbes that are in biofilms. These organisms that are living in it, they attach themselves to this surface. It allows the water to then flow over these microbes and they can kind of acquire the nutrients that are naturally present in the water but yet, they don’t have to float and fight the current. Under a microscope, it would be a cluster of grapes with maybe some type of a matrix or some type of a goo or a slime that might kind of hold those cells together.
Chris - What was the question about biofilms that you were looking at here?
Jeff - Well, it was kind of a serendipitous discovery. But really, what we’re interested in is how organisms make a shift between aerobic lifestyles and anaerobic lifestyles. Meaning, in the presence of oxygen, or living in the absence of oxygen. What Ameya discovered is just that when these organisms that we study grow in the absence of oxygen, they form very robust biofilms. And then we try to extrapolate that as to what's going on with the physiology of this organism or what it might be doing or having this behaviour.
Chris - Ameya, what did you actually do to study this?
Ameya - I looked to see how this specific bacterium called Staphylococcus aureus – typically, it lives on the human body and it doesn’t cause us infection. But under certain conditions, it can cause infection. People have found that its ability to cause infection is related to its ability to form these complex communities called biofilms. So what I looked at is how Staph. aureus sticks to a surface and forms these multicellular communities called as biofilms.
Chris - So how do they go from being an individual free living bacterium to ‘wanting’ to form this community coated by this biofilm and what is it made of?
Ameya - It turns out, when you remove oxygen, it makes it want to go from living as an individual cell to start living as a community. They start to cooperate. A part of this co-operation is that they start producing a sticky glue-like material called as the matrix. This matrix helps these bacteria stick to each other or it helps them stick to surfaces such as human tissues. What happens is the sticky glue-like material, the matrix the bacteria make, it kind of encases the bacteria almost like a raincoat. This protects the bacteria from components of the human immune system so they can come in contact with the bacteria and kill it. This allows the bacteria to cause infections.
Chris - Jeff, what do they make that bacterial raincoat from?
Jeff - Well, I think this is one of the most interesting findings that Ameya made is that it turns out that Staphylococcus aureus in these conditions is actually making this matrix or this glue out of DNA. When you remove oxygen, a certain portion of the population, these cells are killing themselves. As they do that, they break open and they release DNA. As DNA, it turns out as a very sticky material.
Chris - Ameya, do you understand how they get hold of the DNA – the surviving cells – and then use it?
Ameya - That part is not entirely clear yet. However, what is fairly evident is that it does help them stick together in some manner. One idea is that in addition to DNA, the matrix is also made up of proteins. DNA is negatively charge and a lot of the proteins are positively charged. It’s possible that they help kind of form like an electrostatic net which then helps the cells, the proteins, and the DNA all come together, and make those matrix.
Chris - Jeff, we’ve heard that the bacteria kill themselves but is that choice or is it that some randomly die and then their corpses get assimilated by the survivors or is there actually a predestined, “You're going to die and this is how we’re going to kill you by the rest of the population.”?
Jeff - I think that’s a huge outstanding question. We’ve found systems that are actually able to respond to oxygen. We found that certain cells break open, and we found that DNA is released. Now, how certain cells are chosen to have this behaviour is still a huge mystery to us. We’re not exactly sure what the stimuli might be that say, “Okay, you cell break open and you cell form the biofilm and survive, and take advantage of this lysing cell.” But that is going to be one of the questions that we were going to try to address as we move this project forward.
Chris - Is it just bacterial DNA they’ll assimilate because obviously, one of the mechanisms, one of the virulence factors of organisms – gram positives like Staph. aureus is that they lyse our cells and release various micronutrients – iron and so on – that they can then use? But inevitably, there will be DNA coming out in the process. Could they use that too?
Jeff - It’s an excellent question. I'm going to have to say yes. Other researchers in the field have found that Staph. aureus does produce a series of proteins that have the ability to bind DNA. That DNA that they bind is not specific to the DNA of the organism that's producing these proteins. So I guess I would answer the question by saying, “Yes, Staph. aureus could take advantage of human DNA after it’s lysed to those cells to help facilitate the formation of these multicellular communities.
Chris - Ameya lastly, if you can get a handle on how this works, and you can disrupt it, does this mean we potentially have a new way to combat Staph. aureus?
Ameya - Definitely, that’s our hope. In the forthcoming project that we’re looking at, we’re trying to understand how we can take different molecules and make these biofilms dissolve and let antibiotics treat the infection. So, our hope is that by understanding how this biofilm structure occurs, we should then be able to try and figure out a way to treat these infections.
Chris - Jeff…
Ameya - Yes, I would agree with everything that Ameya said. One thing that he didn’t point out though is that Ameya actually discovered the protein that has the ability to sense the presence or absence of oxygen. Now, knowing the signal for that regulatory system, we might be able to go through – work with some chemists, design some drugs – to inhibit this protein that senses either the presence or the absence of this bacterium breathing.
08:01 - Making medicines in yeast
Making medicines in yeast
with Irini Pateraki, University of Copenhagen.
A significant proportion of the drugs in the average doctor’s bag have their roots in nature. Digoxin, artemisin, morphine and salicylates are all good examples. Another is forskolin. This is an incredibly useful therapeutic chemical but it’s also an almost impossibly difficult molecule to make, so doctors have been forced to rely, so far, on a natural source, limiting supply and driving up costs. Now Irini Pateraki has found a way to make yeast brew it up for her, as Chris Smith found out...
Irini - Forskolin is a compound that is produced only from one plant. The plant is called Coleus forskohlii, mainly growing in India, and south Asia. Forskolin is important because it has a lot of pharmaceutical properties that all of them depend on the ability that forskolin has to help in relaxing the vessels of our body. For example, forskolin helps treating hypertension, asthma, glaucoma in the eyes. Actually, it’s one of the very few drugs that are efficient in treating glaucoma. Today, we have in the market medicines that contain forskolin for that purposes.
Chris - But because it comes from one single plant source, we’re therefore dependent on that plant to extract the precursors and the chemical.
Irini - Exactly. Also, we have to have in mind, this plant produces forskolin in relatively low amounts. So, when we need forskolin to give it to the market, this source is not enough. And also, forskolin in that plant is produced in combination with many different metabolites. So this plant is producing something like 70 different metabolites, but quite similar with forskolin. So it’s very challenging to isolate and purify forskolin from this plant.
Chris - Why can't we just make forskolin artificially just in the test tube?
Irini - Because Forskolin is a quite complicated molecule so it’s very challenging chemically to produce it. Also, it will be very expensive. The amounts that you can achieve are very low... so the price and the efficiency is not really favourable.
Chris - On the other hand, if you can work out how the plant is doing it and then steal the genetic recipe then you could recreate the synthetic pathway in another organism and mass produce it.
Irini - This is exactly what we are doing. We went back to the plant, sequenced the transcriptome which means we sequence genes that they were expressed in the plant, and we found out the genes and enzymes responsible for the production of forskolin.
Chris - Did you focus on the part of the plant which makes the highest concentrations of forskolin so that you knew you had a reasonable chance of the genes which are switched on there are the ones which are directly linked to making those chemicals?
Irini - Exactly. This is what exactly we did. So we knew that forskolin generally is produced in the roots and we knew that from the tradition because I mean as I said, forskolin is used for many years. So people knew that they can find it in the roots. But when we started to work on that and we went closer, we found out that the forskolin is not generally produced in the whole root but is produced very specifically in the cork of the root. And then we found out that these cork cells, they contain some oil bodies where these oil bodies, they are able to store forskolin and also forskolin precursors. From this data, we realised that forskolin is also produced in these cells, not only accumulating but also produced. So we went back and we sequenced this very, very specific tissue, and this was a big help for the identification of these enzymes.
Chris - How many enzymes are involved in the whole pathway?
Irini - We found nine different enzymes that were necessary to produce this compound. The difficult part though is to make all these genes to express together and to work together in a different organism to produce what you want them to produce because the environment of a plant is very different than an environment of a yeast cell. A plant cell is much more complicated than a yeast cell so you have to make some adaptations and you have to be sure that all these enzymes that you move from the plant to the yeast cell, you can make them functional.
Chris - Does the yeast tolerate making forskolin okay or is it toxic?
Irini - You could say that it’s toxic but because it’s toxic, here, they have some nice machinery that excrete forskolin out of their cell to the medium.
Chris - How much forskolin does it make?
Irini - In this paper, we have said that it can produce up to 40 mg of forskolin per litre of yeast culture. Actually today, we have managed to get much higher titers.
Chris - How does that compare then in terms of the amount of plant matter you would need to extract the same quantity of the drug? Is this a viable and scalable way of producing forskolin?
Irini - This is totally scalable because of course, you can use as many yeast culture as you like. So you can have several bioreactors while for the plant, the plant needs a whole year to grow and then you have to chop it off to take the roots. When you rely on plants to produce forskolin, you have to depend on yearly growth.
13:60 - Is there gender bias in peer review?
Is there gender bias in peer review?
with Markus Helmer, Yale University
When a scientific paper is submitted for publication, an editor at the journal will usually select - a number of authority figures in the relevant field and ask them to “peer review” the manuscript. So, who gets picked to do the job? Specifically, are male scientists more likely to be asked compared with women? Markus Helmer let Chris Smith know about the issue...
Markus - We were looking into, if there is any kind of bias in the way that editors or reviewers have chosen. In our specific case, we were looking into if gender matters for how reviewers are chosen. We use one specific series of journals, the Frontiers series of journals. They do publish alongside each paper who was the reviewer of that paper and who was the editor of that paper. They have been existing since 2007 and over the years, they have published around 40,000 articles. These 40,000 articles, they were handled by 9,000 editors and around 43,000 reviewers. So altogether, that gives a huge dataset in which we can study the behaviour in the peer review process and look for potential bias.
Chris - How did you interrogate that dataset? What questions did you ask of it?
Markus - We were interested in, if women are unrepresented beyond expectation. It’s a well-known fact that there are just less women than men in science. If there are less women than expected, given that just numerical under-representation and that seems to be the case. We saw that for authors and reviewers while for the editors, the actual number was within the expectation.
Chris - What was the other question you asked?
Markus - The other questions we asked was if there's any bias in the way editors select the reviewers. What we found is that it seemed that editors are for both genders, preferentially chose reviewers of their same gender.
Chris - That’s interesting, isn’t it? So we have got a bias but we’ve got both people being biased in favour of their own sex.
Markus - That's true.
Chris - By how much?
Markus - It appeared that this bias is present for both editor and genders, but it appears also that there's a difference in the way female editors prefer their own gender than the way male editors prefer their own gender. It seems that the same gender preference is much more widespread among male editors. Whereas for female editors, it appears to be restricted to relatively few who were then highly preferential.
Chris - Why do you think we’re seeing this because this is what we dub ‘homophile’ – preference for your own gender?
Markus - Yes. So it appears that the same gender preference is something that might be just human nature in a sense. So you see that in small children already, for example, the way small children form friendship networks, in work environments. It really seems to be a widespread pattern of human behaviour.
Chris - Were you surprised by what you found?
Markus - We were surprised by what we found, yeah. We did expect to find that women are underrepresented but on top of that underrepresentation, there is also the same gender preference. We did not expect that. This is potentially also important because it seems that the number of women in science increases over the years. Very slowly, but it does increase. In a couple of years, even when numerical equity might be reached between the genders, the same gender preference might still be there. So, by just improving the numerical ratio between men, women, the gender bias as a whole will not go away.
Chris - In other words, the system is a bit broken and people’s approach to the system is wrong and if we top up the system with women to achieve parity, it will still relapse to type if we leave it alone anyway.
Markus - It appears so, yeah. So this same gender preference would have to be tackled on top of the numerical underrepresentation.
18:23 - Death sentence
Death sentence
with Colleen Murphy, Princeton University
Nematode worms use a pheromone to kiSome species exist as hermaphrodites, which means that a single organism can be both functionally male and female at the same time. In the case of the nematode worm C. elegans, though, it’s also useful from time to time to be able to produce offspring that are exclusively male. This is because they can mate with other hermaphrodites to boost genetic diversity. But then you don’t want too many males, because when they mate they shorten the lifespans of their partners, and you don’t want them hanging around competing for resources for longer than they’re needed; so how do you get rid of them? Colleen Murphy has discovered the answer - they kill themselves off with a pheromone to which only males are sensitive, as Chris Smith found out...
Colleen - The species, they are what are called hermaphrodites. So the mothers make their sperm and oocyte so they don’t need males. They only use males in times of stress when for example there are conditions where they might want to increase their genetic diversity by crossing. So they want to produce a lot of males in those cases. When those males mate with the mothers, then 50 percent of their progeny will be male and 50 percent will be female so you have these basically bursts of male production. The hermaphrodites turn out to want to get rid of those males. We discovered they have a very clever way of doing that which is the males themselves produce a pheromone that is actually toxic to themselves.
Chris - So you get this boom and bust population situation. There is genetic and there is environmental pressure to increase the number of males when you want to drive diversity, but at the same time, you don’t want that running out of control. So you hardwire into the system a death programme so that as the density of males goes up, they're secreting this pheromone that kills other males. So they will bring about their own demise.
Colleen - Exactly. It’s a very clever way I think of the hermaphrodites being able to say, “Well, we have these males. They need to be around for a while.” But they're actually going to kill themselves by making this dose-dependent toxic compound. And that will eventually drive the population of the males back down so that resumes the mostly hermaphroditic population.
Chris - How do the males detect that pheromone in the environment then?
Colleen - We assume there's three receptors although the receptor for that male pheromone, I don’t believe is known yet. We certainly don’t know it and we don’t know which neurons detect it, but we know that neurons are involved in this. There's a genetic trick where you can make the neurons of those hermaphrodites males. When we did that experiment, those hermaphrodites both produced the male pheromone and they received male pheromone as a toxin.
Chris - Now, when a male mates with a hermaphrodite, that also has a negative effect on the hermaphrodite. Is it bypassing the neurological detection? Is it the same death signal but it’s just getting in via a different route or is that a totally independent mechanism?
Colleen - That seems to be an independent mechanism. We’ve been able to find that those are due to the transfer of sperm and seminal fluid just by using genetic tricks where the males don’t make either of those but they still make a pheromone. The pheromone in that case doesn’t affect the female. It’s really the presence of this germline ramping up of activity from the sperm and unidentified components in the seminal fluid that makes their hermaphrodite die early.
Chris - Have you contemplated doing the experiment where you just stop them being able to kill off these males like this and then just grow generations of them and see what happens to population fitness, population numbers, and so on?
Colleen - So we’ve dreamed about a lot of these types of experiments. In our hands, it’s been very difficult. These animals have at least 300 progeny each and they have about a thousand if they mate and they quickly grow out of control. So, there are lots of thought experiments that we’ve mulled over but we haven't actually carried out any multigenerational – at least not successfully. We’ve done a lot of modelling to try to figure out, what would happen if you just, for example, inhibit their ability to be fertile or produce progeny, and that’s how we were able to show that prediction would be that they'd go back to mostly hermaphroditic species after several generations. Certainly losing it, we haven't thought so much about that. We’re really mostly interested in trying to figure out the mechanism that’s at work there now. What is the mechanism by which they actually kill the animals because it doesn’t look like any of the other lifespan or longevity pathways we’ve worked on previously?
22:43 - Synchronising brain waves
Synchronising brain waves
with Ines Violante, Imperial College London
When we perform demanding mental tasks the brain often has to engage multiple different regions that are specialised at performing specific functions at the same time and then exchange information between them. The theory goes that these disparate brain areas link up by synchronising the pattern of nerve firing in both; so what would happen, Ines Violante wondered, if she artificially enhanced the degree of synchronisation - would a person’s intellect be boosted? Chris Smith found out what she learnt...
Ines - I'm interested in how different regions of the brain talk to each other. So, if you have any sort of cognitive process, if you have to hold information in memory for a couple of seconds, you need different regions of the brain to communicate, to pass the information between each other. People have associated brain waves to those processes.
Chris - Are you saying then that you’ve got a pattern of brainwaves in one place and a pattern of brainwaves in the other, and the brain matches them up, like syncs them up so they're two drums beating at the same time, and that’s how you achieve the sort of coherence between these two brain areas?
Ines - Exactly. So the idea is that you have millions of neurones in one region of the brain firing up and another region and they reach their sync. If they do that in a certain beat, you'll be able to pass information.
Chris - Now was that just fanciful thinking that it was a nice model that would explain possibly how things worked or was there actually objective data supporting that idea?
Ines - There's actually both empirical data and using invasive recordings, and also other electrophysiology techniques like EEG. And there's also models supporting the same idea.
Chris - So if there's already a reasonable body of evidence supporting this idea, what was it you were seeking to find out about it?
Ines - So, we wanted to find out whether we can use stimulation techniques to manipulate the way that this information flow if we can affect behaviour, and will be even more powerful if we can use imaging at the same time. So, if we can use brain scans at the same time and we can understand what we’re doing to the brain when we manipulate how this information is flowing in the brain.
Chris - So, how did you do it?
Ines - We used a technique called transcranial alternating current stimulation which is a form of non-invasive brain stimulation. We combined this with functional magnetic resonance imaging.
Chris - Does this mean then that you're literally putting electrical current into people’s heads?
Ines - That’s right. So, we did that while people are in the scanner and we measured their brain activities.
Chris - By using alternating current, does that mean that you can then drive the rate at which neurons are firing so you'll get that beat synchronisation? That’s what you're going for between diverse regions of the brain.
Ines - Precisely. So we hope that we are able to manipulate those neurons to oscillate at the frequency that we are imposing.
Chris - So what did you ask the subjects to do in the brain scanner then while you did this to them?
Ines - We asked participants to perform two tasks in the scanner with different levels of difficulty. One of the tasks was quite simple. They just had to press a button that matches the direction of an arrow they saw on the screen. The other task was a bit more difficult. We show them numbers and we asked them to press a button when the number they saw in the screen match a specific sequence.
Chris - So, that latter task, that’s a sort of working memory task. They’ve got to hold something in mind whereas the first task is literally to interpret and react to a task. You don’t have to hold information and share information between different brain areas.
Ines - Precisely, yes.
Chris - One would therefore predict that if having synchronous activity in the brain is boosted then they should improve on a more difficult task when you do that.
Ines - Exactly, yeah. So, that was the prediction that we will be able to manipulate this process when you actually need to engage different areas of the brain. When the process that you need to execute to perform is hard enough, they need different parts of the brain to talk to each other.
Chris - Is that what you saw?
Ines - Yes, that’s what we saw. The stimulation would only influence performance in the difficult task.
Chris - Now obviously, if there is a reinforcing effect, a beneficial effect of stimulating these different brain regions so they synchronise... logically, if you were to throw a spanner in the works and these synchronised different brain areas that should impair performance or at least not help it. Did you try doing that?
Ines - Yes, we did. Behaviourally, we didn’t see a significant effect of this synchronous condition.
Chris - But is that just because the brain is pretty good at doing what it’s doing and had you stressed the system more, you might have seen an effect?
Ines - That might be the case. It might be that if we got to an even harder condition, we will see bigger spread of the effects.
Chris - You had these subjects in the brain scanner while this is going on. You’re either stimulating the brain disparate regions at the same time synchronously or asynchronously so they're offbeat. What do the brain scan show when you do this?
Ines - The synchronous stimulation shows an increase in brain activity in the regions that are involved in working memory. The asynchronous stimulation shows increasing brain activity in regions that are not involved with working memory.
Chris - So that’s sort of logical. You'd expect if you're just recruiting some brain areas, you're going to get a bit of nonspecific brain recruitment which is not related to the task in hand.
Ines - Yes, indeed. But it was quite interesting to actually be able to see those effects and to be able to see that even functional connectivity – a measure of how different areas of the brain are talking to each other – they're also changed.
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