Brainless Intelligence: Memory in Physarum polycephalum

by JH

Slime mold

     “If I only had a brain...”, cooed the Scarecrow from the hit classic, The Wizard of Oz. A brain he longed for so that he could think, retain memories and reflect intelligence, just like Dorothy and the other complex beings he encountered on his journey down the yellow brick road. Thoughts and memory reside in the brain, but what if an organism doesn't have a brain? Can it still possess the qualities that constitute intelligence, such as memory? In the case of the slime mold, Physarum polycephalum, the answer is yes. Scientists have explored the ability of the single-celled protist to construct a form of external memory to navigate through complex environments (1). This behavior demonstrates a primitive version of brain function in P. polycephalum, despite their lack of a central nervous system (3).

     Physarum polycephalum is a unicellular protist of the class Myxomycetae, better known as a slime mold, which is most commonly found in the organic matter of the forest floor such as woody debris and leaf litter (2). P. polycephalum shares the same bright yellow coloring as the popular kid’s television character “SpongeBob SquarePants”. Unlike its famous doppelgänger, P. polycephalum is far from simpleminded. The slime mold enhances its navigational ability through the use of an external spatial memory system, in which the organism can distinguish between new and previously explored pathways. The purpose of this “memory” is to determine the most efficient and direct route to a food source as well as prevent the organism from revisiting areas of depleted nutrients (1). This ability to construct a spatial memory system even allows P. polycephalum to find its way through a complex maze!

     How does this organism achieve such intelligible feats? Researchers studying Physarum polycephalum believed this organism may use a type of avoidance technique to forgo areas of previous exploration in order to find the most efficient route to a food source. They hypothesized that the slime mold uses the presence of extracellular slime, which is a thick layer of cells secreted by the organism, as a spatial memory system to aid in this avoidance behavior (1). To observe the behavior of P. polycephalum, a team of Australian scientists set up an experiment modeling a classic test of navigation that is commonly used in robotics, called the U-shaped trap problem (1). This test required the slime mold, in its vegetative state called plasmodium, to reach a goal behind a U-shaped barrier. The test consisted of a petri dish with a U-shaped trap, made of a material that the organism could not stick to and grow over, that was placed between the slime mold and the goal, a solution of sugar. The team challenged the slime mold to reach the sugar solution by growing around the trap on two types of media: blank agar and agar containing extracellular slime.

     What they found was that the slime mold strongly avoids areas that contain the extracellular slime (1). They observed that the organism first explores its entire environment by extending temporary projections that help the organism move, known as pseudopodia. The organism then detects where the food source is via specific receptor molecules. This detection causes the projections to flow towards the food source while simultaneously retracting the pseudopodia from all of the areas that do not contain food (1). As the plasmodium retracts its pseudopodia, it leaves behind a thick extracellular slime. It can then use this slime as a memory system by “reminding” the organism upon contact with the slime, that these areas did not contain food. The slime mold then grows exclusively along the shortest path avoiding all of the areas containing extracellular slime.

     This behavior was also observed in an experiment where P. polycephalum was placed at the start of a complex maze, as shown in Fig.1 (3). Food sources were placed at the starting and ending points of the maze, and the slime mold was encouraged to find a route that leads to the food source at the end. The observation was that the organism extended its pseudopodia throughout all possible paths of the maze, even the dead ends. Once the organism had completely covered the maze, it began to retract its pseudopodia, avoiding the slime in the dead end areas, and began to grow only along the path that offered the shortest distance to the food source at the end (3).

Fig.1 Slime mold solving a maze;
finding the shortest distance to food source

    

     Another aspect of this experiment explored the behavior of P. polycephalum when extracellular slime from other organisms of the same species as well as members of different species of slime mold were deposited in its presence. The results of this experiment showed that the organism discriminates between the two groups (2). Because P. polycephalum naturally coexists with a variety of species of slime mold, which compete for similar yet slightly different food sources, this distinction between slime from different organisms is essential for maximizing foraging efficiency (2). A member of the same species of P. polycephalum will have nutrient requirements that overlap entirely therefore the detection of its extracellular slime suggests that the food source is already depleted. The experimental P. polycephalum thus will avoid these areas. The organism would then prefer to follow cues from a member of a different species, as this would indicate an environment of slightly higher quality and less depletion than that of a member of the same species (2).

     Maze solving, network construction, and species specific discrimination are used by the slime mold Physarum polycephalum to allow the organism to more efficiently focus its search for nutrients. The studies previously discussed are important in that they show empirical evidence of a spatial memory system in a non-neuronal organism to make optimal foraging decisions. These conclusions support the theory that externalized memory may be a functional precursor to the internal memory of complex organisms (1). Just like the Scarecrow, Physarum polycephalum proves that even without a brain, you can still find your way to the end of the yellow brick road.


References

1. (1)  Reid, C.R., Latty, T., Dussutour, A., Beekman, M. 2012. Slime Mold Uses an Externalized Spatial “Memory” to Navigate in Complex Environments. PNAS vol. 109 no. 43 http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3491460/pdf/pnas.201215037.pdf
2. (2)  Reid, C.R., Latty, T., Dussutour, A., Beekman, M. 2013. Amoeboid Organism Uses Extracellular Secretions to Make Smart Foraging Decisions. International Society for Behavioral Ecology vol. 24 ch. 4 pgs 812-818 http://beheco.oxfordjournals.org.ezp2.lib.umn.edu/content/24/4/812.full.pdf+html
3. (3)  Nakagaki, T., Yamada, H., Tóth, A. 2000. Maze Solving By an Amoeboid Organism. Nature vol. 407 pg 470 http://www.nature.com.ezp2.lib.umn.edu/nature/journal/v407/n6803/pdf/407470a0.pdf
4. (4)  Nakagaki, T., Kobayashi, R., Nishiura, Y., Ueda1, T. 2004. Obtaining multiple separate food sources: behavioural intelligence in the Physarum plasmodium. The Royal Society http://rspb.royalsocietypublishing.org/content/royprsb/271/1554/2305.full.pdf

Circadian Rhythm Regulation in Chlamydomonas

by BA

How many times have you stayed up late for a party or, more likely but less fun, for last minute work, sacrificing your usual sleep schedule with the thought that “I can sleep when I’m dead”? You probably found that it’s difficult to catch up on that lost sleep, and sometimes it even takes a couple days to get that sleep-wake cycle back to where it should be.

You can thank your circadian rhythm for that. The circadian rhythm is the 24-hour cycle that regulates your sleep patterns and other physiological processes, like blood pressure and hormone levels, based on your environment. It is regulated by your circadian clock, which makes it possible for you to coordinate your behavior with environmental changes that result from the day-night cycle. Light and temperature can affect it, as evidenced by animal migration and seasonal depression in some people, and it can be located in various parts of the organism’s body. For example, monarch butterflies have circadian clocks in their antenna (1), plants have cells that detect light (known as photoreceptors) to help regulate certain genes, and humans have circadian clocks in a tiny region of the brain called the suprachiasmatic nuclei. Interestingly, unicellular organisms can have circadian clocks too, such as the photosynthetic green algae Chlamydomonas. Why do these little guys need a circadian rhythm, and how have studies on them helped us understand our own?

Chlamydomonas cell with its
components labeled. (8)    

Chlamydomonas has a small, light-sensitive organelle near the middle of the cell known as an eyespot that allows it to move depending on the intensity of the light in its environment. Cells like Chlamydomonas are attracted to sources of light in a phenomenon we call positive phototaxis. (2) This allows cells to find light sources during the day so they can utilize their photosynthetic mechanisms more efficiently. (3) When the night comes and very little light is to be found in the environment, the cell turns to chemotaxis, which is the movement towards or away from certain chemicals or molecules. Chemotaxis allows the cells to find sources of nitrogen, an essential nutrient for photosynthetic organisms. (4) This type of switching behavior allows Chlamydomonas to use its energy in the most efficient way possible; trying to use photosynthesis at night would be useless due to the lack of sunlight, and using chemotaxis to find nitrogen during the day would only waste precious time that would be better spent on photosynthesis. The constant rotation between phototaxis during daylight and chemotaxis at night is indicative of circadian rhythms in the organism, which means there must be a circadian clock. Indeed, many genes for the circadian clock in Chlamydomonas have been identified, including gene families (sets of genes with a similar functions) responsible for phototaxis, and several independent genes attributed to the overall robustness of the circadian clock mechanism. (5)

One gene family in Chlamydomonas called per has been identified as a main regulator of circadian clock periods. Chlamydomonas cells with mutant per genes, wherein the DNA of the genes has been somehow damaged, have been demonstrated to have shortened or lengthened clock periods. (5) By allowing cells with mutant per to grow in culture, waiting an expected number of normal circadian cycles, and testing phototaxis, scientists were able to determine how the mutant genes disrupted the circadian rhythm of the organism. If some cells underwent phototaxis when they weren’t expected to, like during times without sunlight, they were presumed to be mutants. This was confirmed with genetic experiments that involved allowing the mutants to sexually reproduce and examining the new cells’ phototactic movements, which further revealed the existence of multiple genes responsible for the mutants. (5)

Another gene, CK1, encodes a protein kinase, which is an enzyme that changes the function of other proteins by adding phosphate groups to them. CK1 is known to be one of the proteins specific to both the Chlamydomonas eyespot and flagella, a whip-like organelle used for locomotion. (6) The presence of CK1 in both the light-sensing and movement organelles implies that it is important to phototaxis. To test this idea, the functional CK1 gene was prevented from working in the cell, known as knocking out the gene, by a method called RNA interference where RNA was used to specifically block the gene from being expressed. This interference resulted in defects in the formation of flagella, where cells synthesized small flagella or even none at all. (6) RNA interference can be made to be stronger or weaker, and in this case, as interference got stronger, more severe defects in flagella were observed. These CK1 knockout cells were also subjected to phototaxis tests where they were exposed to bright light to mimic the day and darkness to mimic the night. While cells with the normal CK1 gene (wild-type cells) exhibited normal attraction to light during the day and no attraction during the night, the CK1 knockout cells showed significantly lower attraction to the light in the daytime. (6) This can be described by the mutation of the CK1 gene not being able to express its protein kinase. Without the CK1 protein in the Chlamydomonas cells, the eyespot was less able to detect light, and deformed flagella was not sufficient to move the cell towards whatever light it could detect.

ROC genes are suspected to be core genes of the Chlamydomonas circadian clock. They encode transcription factors, which control the expression of other genes in the journey from DNA to proteins. (7) Cells with mutations in their ROC genes display strange circadian cycles, either short, long, or inconsistent. Because these genes control expression, scientists thought they might have a role in feedback mechanisms that regulate the circadian clock, mechanisms in which the product of gene expression determines whether more product is made or not. (3) In particular, two proteins, ROC75 and ROC40, were suspected to have some sort of interaction. This was examined by fusing fluorescent tags to each protein and watching where they went in the cell. T. Matsuo presented his findings at the 14th Annual Chlamydomonas Conference in 2010, explaining that ROC75 was found in around the nucleus of the cell and interacted with the promoter region of the ROC40 gene, part of the DNA that controls the initiation of gene expression. This suggests that ROC40 expression is regulated by ROC75. Levels of expressed ROC genes were then shown to change depending on the phase of the circadian cycle (3). This evidence tells us that genes with feedback mechanisms that respond to day-night cycles form the basis of the circadian clock in Chlamydomonas.

The cool thing is that these genes have homologs in other eukaryotes, including us. (3) Chlamydomonas circadian clock genes have strong homology (similarity) with genes found in both plants and higher animals. Some genes in the ROC family share similarities with the genes responsible for the circadian clock and seasonal flowering in the plant Arabidopsis thaliana. (3) As another example, CHLAMY1, a protein in Chlamydomonas that binds to RNA, has a subunit named C3 that is extremely similar to the protein CUGBP2 in rats. In fact, if an antibody that specifically targets the Chlamydomonas C3 protein is introduced into rats, it will recognize the CUGBP2 protein in the suprachiasmatic nuclei of the rat brain – the same part of the brain that is responsible for the human circadian rhythm. This result suggests numerous similarities between higher animals and the unicellular Chlamydomonas, making it a good model for examination of the human circadian rhythm.

The circadian rhythm allows us to complete our day-to-day tasks as best we can, both conscious (working our jobs and sleeping the proper amount each night) and unconscious (hormone regulation and tissue repair). While Chlamydomonas doesn’t use it for sleeping like humans, it requires functional circadian clock genes to make efficient use of its energy and thrive. By studying this little algae, we can better understand ourselves and the world around us.

References

1. Merlin C, Gegear RJ, Reppert SM. 2009. Antennal circadian clocks coordinate sun compass orientation in migratory monarch butterflies. Science. 325(5948): 1700-1704.
2. Stavis RL, Hirschberg R. 1973. Phototaxis in Chlamydomonas reinhardtii. J Cell Biol. 59(2): 367-377.
3. Matsuo T, Ishiura M. 2011. Chlamydomonas reinhardtii as a new model system for studying the molecular basis of the circadian clock. FEBS Lett. 585(10): 1495-1502.
4. Fernandez E, Galvan A. 2007. Inorganic nitrogen assimilation in Chlamydomonas. J Exp Bot. 58(9): 2279-2287.
5. Matsuo T, Ishiura M. 2010. New insights into the circadian clock in Chlamydomonas. Int Rev Cell Mol Biol. 280: 281-314.
6. Schmidt M, Gessner G, Luff M, Heiland I, Wagner V, Kaminski M, Geimer S, Eitzinger N, Reissenweber T, Voytsekh O, Fiedler M, Mittag M, Kreimer G. 2006. Proteomic analysis of the eyespot of Chlamydomonas reinhardtii provides novel insights into its components and tactic movements. Plant Cell. 18(8): 1908-1930.
7. Brunner M, Merrow M. 2008. The green yeast uses its plant-like clock to regulate its animal-like tail. Genes & Dev. 22:825-831.
8. Chlamydomonas photo courtesy of Connecticut College. Accessed November 12, 2015. http://fmp.conncoll.edu/Silicasecchidisk/LucidKeys3.5/Keys_v3.5/Carolina35_Key/Media/Html/Chlamydomonas_Main.html

A Different Kind of Red Menace

by JR

Manatee Killing Red Tide

The idea of large areas of the ocean running red sounds like something out of a horror movie, as though Jaws were a cautionary documentary about bloodthirsty sharks instead of the original summer blockbuster. Instead, red waters are associated with an increasingly common phenomenon called a red tide or, more accurately, a harmful algal bloom (HAB). Harmful algal blooms in the Gulf of Mexico off the west coast of Florida are caused by the photosynthetic unicellular organism Karenia brevis, a type of dinoflagellate or marine plankton. While hardly the type of organism to inspire a week-long celebration on the Discovery Channel, the fact remains that K. brevis harmful algal blooms cause many problems. These HABs adversely affect animals and humans that are exposed to such blooms in Florida.

HABs of K. brevis are thought to begin by upwelling of nutrients promoting the growth of K. brevis near the surface of coastal waters [1]. What makes K. brevis so problematic is the organism’s ability to produce a unique type of neurotoxin, called brevetoxins. These brevetoxins are released when the cell is destroyed by factors such as wind or waves [2]. During a HAB event, the high concentrations of K. brevis cells make this release of brevetoxins a problem for marine species, as well as humans living near the coast. Even after a HAB event has ended, brevetoxins can remain in the water and later be released into the air through bubbles that rise to the surface, where they are incorporated into marine aerosols that can travel inland [2]. In a study done by Hardison et al, it was shown that K. brevis produced more brevetoxins under phosphorous (P) and nitrogen (N) limited conditions [1]. The production of brevetoxins in P-limited conditions was twice that of N-limited conditions on top of the observed increase in both conditions, prompting a need to measure the amount of brevetoxins rather than just the concentration of K. brevis [1].

K. brevis

Florida is a popular tourist destination, so HABs can negatively impact the Florida tourist industry, as red waters and the risk of negative effects from the toxins are hardly a draw for tourists. This has negative economic impacts in areas that depend on tourism and fishing, as, for example, the tourism industry in Florida is worth $57 billion [3]. In humans, one such negative effect is that inhaling airborne brevetoxins can cause respiratory irritation, such as coughs or bronchitis, the effects of which are more potent and therefore measurable in people who have asthma [3]. In a study done by Hoaglund et al, the costs of the respiratory illnesses were estimated to be $0.5-$4 million in Sarasota County, Florida alone, depending on severity [2].

Another human risk factor associated with brevetoxins is neurotoxic shellfish poisoning (NSP). Commercially, shellfish are monitored for brevetoxins, but this does not stop people from recreationally collecting and eating shellfish that might contain brevetoxins [4]. Shellfish are filter feeders, so they can consume the K. brevis cells during a HAB event, leading to the buildup of brevetoxins in shellfish tissues [5]. When these shellfish are eaten by humans, the brevetoxins can cause symptoms associated with NSP ranging from stomach upset to partial paralysis [4]. No deaths have been associated with either factor, but they are a cause for alarm nonetheless [2, 4].

Humans are not the only creatures affected by these HAB events. Fish kills in Florida are most commonly caused by HABs—a shocking 96% of them between 2003 and 2007 [6]. Fish can absorb brevetoxins across their gill membranes, or by consuming other fish that have similarly been exposed to the toxins, leading to the death of the fish [7]. These fish kills might seem advantageous to shore birds and other marine animals that feed on the fish, but brevetoxins can travel up the food chain. Furthermore, the presence of large amounts of dead fish choke up the water, resulting in a lack of oxygen for other species that live there [7].

As mentioned, brevetoxins present in living or dead fish can travel up the food chain. Van Deventer et al observed shore birds feeding on HAB-killed fish on beaches in central west Florida [8]. The researchers collected dead fish and tested them for brevetoxins, and they also collected shore bird carcasses to similarly test for brevetoxins [8]. The tested fish were found to have levels of brevetoxins that could be potentially lethal to small shore birds, and the shore birds also had brevetoxins present in their digestive tracts [8]. Although the researchers did not conclude that the shore birds died from consumption of fish killed during a HAB event, there is evidence that the fish were indeed the source of the brevetoxins found in the birds [8].

Another example of the dangers of brevetoxins to Florida wildlife can be found in manatees. Exposure to high levels of brevetoxins can be lethal to Florida manatees, but exposure at lower levels may adversely affect their immune systems, as well [9]. In a study by Walsh et al, the researchers took plasma samples from rescued manatees recovering at the Lowry Park Zoo in Tampa, Florida [9]. The researchers tested the plasma samples for brevetoxins, as well as various tests to determine the fitness of the manatees’ immune system [9]. The researchers found a relationship between increased brevetoxin levels and decreased lymphocyte proliferation, suggesting that brevetoxins may have a negative, immunosuppressant effect on the manatee immune system [9]. The Florida manatee is an endangered species, a status aggravated by human activity, and exposure to K. brevis HAB events further threatens a species already at risk.

Red tides caused by K. brevis may not be preventable, but understanding of its effects and monitoring can be useful in mitigating the negative effects of such events. A system piloted in Sarasota County and Manatee County, Florida called Integrated Ocean Observing System allowed lifeguards at eight public beaches to report different conditions on the beach, such as dead fish and respiratory distress of beach goers [3]. The data was then transferred to a server accessible by the public [3]. Similar databases in other areas at risk of HABs would be helpful to limit human exposure to airborne brevetoxins, as well as shellfish that may have accumulated brevetoxins in their tissues. Further, understanding of the risks can help with conservation of animal species at risk of exposure to K. brevis brevetoxins. Red oceans caused by algal blooms might not be as glamorous as waters blood-stained by shark attacks, but unlike uncommon shark attacks, algal blooms pose a common risk that should be no less urgent.

References:

1. Hardison, D.R., Sunda, W.G., Shea, D., and Litaker, R.W. (2013). Increased Toxicity of Karenia brevis during Phosphate Limited Growth: Ecological and Evolutionary Implications. PLoS One 8.

2. Hoagland, P., Jin, D., Polansky, L.Y., Kirkpatrick, B., Kirkpatrick, G., Fleming, L.E., Reich, A., Watkins, S.M., Ullmann, S.G., and Backer, L.C. (2009). The Costs of Respiratory Illnesses Arising from Florida Gulf Coast Karenia brevis Blooms. Environ Health Perspect 117, 1239–1243.

3. Kirkpatrick, B., Currier, R., Nierenberg, K., Reich, A., Backer, L.C., Stumpf, R., Fleming, L., and Kirkpatrick, G. (2008). Florida red tide and human health: A pilot beach conditions reporting system to minimize human exposure. Science of The Total Environment 402, 1–8.

4. Watkins, S.M., Reich, A., Fleming, L.E., and Hammond, R. (2008). Neurotoxic Shellfish Poisoning. Mar Drugs 6, 431–455.

5. Abraham, A., Wang, Y., El Said, K.R., and Plakas, S.M. (2012). Characterization of brevetoxin metabolism in Karenia brevis bloom-exposed clams (Mercenaria sp.) by LC-MS/MS. Toxicon 60, 1030–1040.

6. Gannon, D.P., McCabe, E.J.B., Camilleri, S.A., Gannon, J.G., Brueggen, M.K., Barleycorn, A.A., Palubok, V.I., Kirkpatrick, G.J., and Wells, R.S. (2009). Effects of Karenia brevis harmful algal blooms on nearshore fish communities in southwest Florida. Mar Ecol Prog Ser 378, 171–186.

7. Pierce, R.H., and Henry, M.S. (2008). Harmful algal toxins of the Florida red tide (Karenia brevis): natural chemical stressors in South Florida coastal ecosystems. Ecotoxicology 17, 623–631.

8. Van Deventer, M., Atwood, K., Vargo, G.A., Flewelling, L.J., Landsberg, J.H., Naar, J.P., and Stanek, D. (2012). Karenia brevis red tides and brevetoxin-contaminated fish: a high risk factor for Florida’s scavenging shorebirds? Botanica Marina 55, 31–37.

9. Walsh, C.J., Butawan, M., Yordy, J., Ball, R., Flewelling, L., de Wit, M., and Bonde, R.K. (2015). Sublethal red tide toxin exposure in free-ranging manatees (Trichechus manatus) affects the immune system through reduced lymphocyte proliferation responses, inflammation, and oxidative stress. Aquatic Toxicology 161, 73–84.

The mitochondrion: where did the powerhouse of the cell come from?

By Scott Miller

Nestled along the Minnesota River within view of the freeway, Black Dog Generating Station is an Xcel Energy power plant with a capacity of around 500 megawatts in Burnsville, Minnesota. Built in the 1950s, Black Dog generates power from coal and gas.1 As a child, I remember driving by this behemoth, seeing black curls of pollution rising into the sky. To me, it had always been a fixture along the river, but one day I questioned how this powerhouse got there. Biologists have had the same question about mitochondria.

Living organisms are broadly grouped into two camps: prokaryotes and eukaryotes. Prokaryotes encompass bacteria and another group of microbes called archaea, while eukaryotes include many microbes but also more complex organisms such as plants, fungi, and animals. Eukaryotes and prokaryotes differ in many ways, but eukaryotes generally have a membrane-bound nucleus, whereas prokaryotes do not.2 In addition to a nucleus, eukaryotic cells can have other membrane-bound organelles as well, though precisely which organelles the cells have depends on the species. Besides the nucleus, another common organelle is the mitochondrion. If there is one thing students remember from high school biology, it is that “mitochondria are the powerhouses of the cell.” This is (broadly) true, but most high school biology classes stop short of following up with a much more exciting lesson: where did mitochondria come from?

Biologists view life through an evolutionary lens. Novelties do not spontaneously arise; they have to come from something. Eukaryotes did not just suddenly sprout functioning mitochondria. Consequently, evolutionary biologists have developed a widely accepted theory for how these cells acquired their “powerhouses,” termed the endosymbiotic theory.3 According to the endosymbiotic theory, at some point in distant evolutionary history, one larger cell engulfed a smaller cell. The smaller cell continued to live inside the larger cell and perform functions similar to today’s mitochondria. Over generations and with evolution, the smaller cell lost its autonomy, evolving into the mitochondrion for the larger cell.

Mitochondria in bovine lymphocytes. 

The features of mitochondria support endosymbiotic theory in that they share many characteristics with prokaryotes, the likely “smaller cell.”4 First, mitochondria are roughly rod-shaped, reminiscent of the structure and size of many prokaryotes. The second feature is replication. In humans, for example, most cells replicate in a process called mitosis. Mitochondria, however, divide independently of mitosis in a process similar to binary fission, which is how bacteria replicate.5 Another feature of mitochondria supporting endosymbiotic theory is the presence of DNA inside mitochondria. Mitochondrial DNA is separate from the DNA found in a eukaryote’s nucleus and has a circular structure similar to DNA found in prokaryotes.

Biologists wanted to see if any existing species of eukaryotes and prokaryotes resembled an early mitochondrion and its larger host cell. Using modern sequencing techniques, biologists have been able to determine the combination and order of nucleotide bases that make up the DNA in mitochondria. Biologists have also been able to sequence the DNA from many prokaryotes. They can then compare the DNA from mitochondria and the DNA from prokaryotes. The idea is that similar DNA sequences correspond to a mitochondrion and prokaryotic species that are more closely related to each other.

Researchers consider the small freshwater eukaryote Reclimonas americana to have the most ancestral mitochondria; that is, the mitochondria in R. americana are the most similar to bacteria of any eukaryote. R. americana is part of a larger group of species called jakobids. Not all eukaryotes have mitochondria—though all eukaryotes have at least one nucleus—and jakobids are very similar to retortamonads, a different group of eukaryotes lacking mitochondria. A group of scientists from Canada and New England therefore hypothesized that jakobids represent a point in evolutionary history where eukaryotes first acquired mitochondria.6

 To test their hypothesis, these scientists examined the sequence of the DNA from R. americana’s mitochondria. The researchers found 92 functional genes in R. americana mitochondrial DNA, compared to only 37 in DNA from human mitochondria. The 92 genes included the code for several proteins normally found in bacteria. Of the 67 proteins that R. americana mitochondria can make with their DNA, 18 are not found in other eukaryotes’ mitochondria. Most notably, the researchers identified four rpo genes in R. americana, all of which had never been found in mitochondria prior to this study. The rpo genes contain the code for proteins called RNA polymerases, which play a prominent role in the biological process of transcription. The rpo genes found in R. americana’s mitochondria are usually only found in bacteria.

The researchers also identified other key features linking R. americana mitochondria to bacteria. One was organization of the DNA in the mitochondria. Bacterial DNA often contains operons, which are clusters of genes that have products with a similar function. For example, all of the genes coding for proteins involved in digesting lactose reside in the lac operon. The mitochondrial DNA in R. americana contains vestiges of operons. For example, the rpo genes were all linked together in the mitochondrial DNA. This organization provides further proof of a relatively close relation between R. americana mitochondria and bacterial cells.

Additionally, the researchers found evidence for Shine-Dalgarno-type sequences in R. americana mitochondrial DNA. Without elaborating on the technical details, Shine-Dalgarno sequences are normally a distinct characteristic of prokaryotes. If they are present in R. americana mitochondria, this constitutes additional evidence that these mitochondria are relatively closely related to prokaryotes.

In addition to finding which mitochondria are most related to prokaryotes, biologists have studied which prokaryotes are most similar to mitochondria. A group of French scientists compared the mitochondrial DNA sequence from R. americana to those of several bacterial species from the grouping known as the Rickettsiales order.7 Scientists have suspected that the bacterial ancestor of mitochondria came from the Rickettsiales. The French scientists analyzed the DNA sequence of nine genes found in both the bacteria and R. americana mitochondria. Using statistics and software, the researchers confirmed that the bacterial ancestor of mitochondria was related to other Rickettsiales. In particular, they determined that the bacterial species Pelagibacter ubique is most similar to R. americana mitochondria.

Research studies continue to support endosymbiotic theory for the origin of mitochondria. The current thought is that Reclimonas americana mitochondria are the most closely related to bacteria of any mitochondria. In turn, the bacterial species Pelagibacter ubique most closely resembles the bacteria that developed into mitochondria. While the picture is not entirely clear, biologists now have a much better idea of where mitochondria came from.

Works Cited

1. Black Dog Generating Station, 2015. Available from: . [10 December 2015].
2. John, P., and Bell, L. (2001). Viral Eukaryogenesis: Was the Ancestor of the Nucleus a Complex DNA Virus? Journal of Molecular Evolution 53(3), 251-256.
3. Davidow, Y., Huchon, D., Koval, S.F., and Jurkevitch, E. (2006). A new α-proteobacterial clade of Bdellovribrio-like predators: implications for the mitochondrial endosymbiotic theory. Environmental Microbiology 8(12), 2179-2188.
4. Gray, M.W., Burger, G., and Lang, B.F. (2001). The origin and early evolution of mitochondria. Genome Biol 2, reviews1018.1–reviews1018.5.
5. Seo, A.Y., Joseph, A.-M., Dutta, D., Hwang, J.C.Y., Aris, J.P., and Leeuwenburgh, C. (2010). New insights into the role of mitochondria in aging: mitochondrial dynamics and more. J Cell Sci 123, 2533–2542.
6. Lang, B.F., Burger, G., O’Kelly, C.J., Cedergren, R., Golding, G.B., Lemieux, C., Sankoff, D., Turmel, M., and Gray, M.W. (1997). An ancestral mitochondrial DNA resembling a eubacterial genome in miniature. Nature 387, 493–497.
7. Georgiades, K., Madoui, M.-A., Le, P., Robert, C., and Raoult, D. (2011). Phylogenomic Analysis of Odyssella thessalonicensis Fortifies the Common Origin of Rickettsiales, Pelagibacter ubique and Reclimonas americana Mitochondrion. PLoS ONE 6, e24857.