1Name: JohnathanTsui Wei YauCourse: Microbiology and VirologyStudent number: 1222880Date:10-04-2013Word count: 2964Extended essay: Reviewing the evidence for the Endosymbiotic theory-the modern, or organelle containing eukaryotic cell evolved in stepsthrough the stable incorporation of chemo-organotrophic andphototrophic symbionts from the domain BacteriaAbstract (100 words max): An introduction to the Endosymbiotic theory is first given toprovide an overview of the content of discussion, more specifically the evidence and theories for andagainst it. The next two sections serve to describe the two main alternative theories as opposed to theSET, preventing the assumption that it is the most validated theory. Next section that ensues discussesmainly evidences which support the SET,it is further divided into two sub-sections using two of themain eukaryotic organelles concerned- mitochondria and chloroplasts. Finally, a conclusion regardingall the surrounding arguments discussed is given.
2A brief introduction to Endosymbiotic theoryTo put it bluntly, Endosymbiotic theory (or SET, Serial Endosymbiosis Theory) is a theory that tries toexplain the evolutionary origin of organelles in modern day eukaryotic cells. “As early as 1883, A.F.WSchimper proposed that the chloroplasts of plants were cyanobacteria that lived symbiotically insideplant cells…..Early scientists such as Ivan Wallin were convinced that mitochondria were bacteria thatlived inside cells.” (Paracer and Ahmadjian 81) It was first revitalized and put forth in the 1960’s byLynn Margulis, a biologist from Boston University, and published in her book “Symbiosis in CellEvolution” in 1981. It proposes that organelles residing in eukaryotic cells, most notably mitochondriaand chloroplasts (possibly peroxisomes, hydrogenosomes, etc other membrane bound organelles),originated from the endocytosis of aerobic eubacteria and cyanobacteria (respectively) by anaerobicprokaryotic host cells (archaebacteria). Unable to digest the engulfed bacteria, they developed asymbiotic relationship (where two species gain mutual benefits by living alongside each other) overbillions of years, hence the name “endosymbiosis”. The prokaryotes that were once unable to utilizeoxygen began to acquire abilities to carry out aerobic respirations and eventually photosyntheticreactions by successive endosymbiosis events. As time progressed, the engulfed bacteria transferredsome genes to the host cell’s genome, resulting in the permanent interdependence on each other’smetabolic activities. Eventually, they developed into some of the known organelles such asmitochondria and chloroplasts.
3As a boldly proposed theory, the SET was questioned by the scientific community throughout the 19thand 20thcentury as there is no way to support it with experimental results. However, the advancementof scientific knowledge and technologies in recent years allows the structural, molecular andbiochemical scrutiny of eukaryotic organelles to construct probable backup on the SET. Some of theseevidences include the fact that both mitochondria and chloroplasts possess their own circular DNA(though not all in mitochondria), which strongly resembles prokaryotic cells; the fact that theseorganelles synthesize their own proteins and perform binary fission is also a powerful evidence.Some of the evidences which hint the validity of SET will be examined in the following sections of thisessay, while arguments that the SET cannot be established will also be examined in order to provide anobjective, bird eyes’ view on this particular subject. Before this, however, some of the other theoriesand hypothesis regarding the rise of eukaryotic organelles will also be mentioned for the sake ofbalancing alternative views on this matter.
4Autogenous TheoryFigure 1- this is a simple illustration of how infoldings of an ancestral prokaryote can result in single membraneorganelles such as ER and nucleus, which leads to further compartmentalization of plasmids within theseinvaginations that results in the appearance of organelles like mitochondriaThe autogenous theory proposes that organelles such as nucleus, endoplasmic reticulum, Golgiapparatus, vacuoles and lysosomes first “evolved from elaborations of the cell and other internalmembranes” (Paracer and Ahmadjian 80), or in other words progressive compartmentalization as seenin figure 1. These single membrane organelles arise from the infoldings of a prokaryotic ancestor cell’splasma membrane (Gwu.edu), and organelles such as mitochondria and chloroplasts successivelyevolved after the appearance of a prototype eukaryote by compartmentalization of plasmids within theinvaginations of the cell membrane (Gwu.edu). The common features found between mitochondria &chloroplasts and eubacteria as proposed in SET might be explained by mosaic evolution, where “thecomponents in the compartment evolved more slowly than other parts of the cell” (Gwu.edu), thusexplaining why they possess some of the molecular and structural similarities to a prokaryotic cell. The
5fact that they have double membrane can also be explained by possible, secondary invaginations of theplasma membrane.The Hydrogen HypothesisFigure 2- On the left hand side, type II amitochondriate eukaryote resembles the anaerobic proteobacterium thatalso utilizes hydrogenosome to generate ATP, with hydrogen, carbon dioxide and acetate discarded as wasteproducts; this proposed endosymbiont is compared to a eukaryote with mitochondria as seen on the right
6Figure 3- this is a simple illustration of how the archaen (methanogen), which takes in hydrogen, carbon dioxideand acetate to produce ATP with methane discarded as waste products, engulfed the proteobacterium (hydrogen-producing symbiont) over timeIn short, the hydrogen hypothesis “is based on the symbiosis between an alpha proteobacterium and anarchaean (methanogen)” (Paracer and Ahmadjian 81). In aerobic cell respiration performed bymitochondria (following glycolysis), pyruvate is decarboxylated into acetyl-CoA by the PDF (Pyruvatedehydrogenase complex), which is then oxidized in the citric acid cycle into NADH, CO2 and water,together with 36 mol of ATP for each mol of glucose consumed. The proteobacterium involved in thesymbiosis performed respiration similar to an anaerobic amitochondriate eukaryote withhydrogenosome, where pyruvate is converted into acetate, CO2 and reduced ferredoxin by the pyruvateferrodoxinoxidoreductase (PFO). The reduced ferredoxin is reoxidized by hydrogenase, producing ATPand hydrogen, hydrogen is discharged with carbon dioxide and acetate as waste products, as seen infigure 2. On the other hand, the archaean involved are methanogens- obligate anaerobe that useshydrogen and carbon dioxide to produce ATP (Acetyl-CoA pathway) and methane (Methanogenesis),
7with methane discarded as waste products. This ensures that symbiosis took place as mutual benefitsbetween these two organisms were guaranteed. The archaean became the host with the proteobacteriumas the endosymbiont, as depicted in figure 3. Eventually, the host became dependent on theproteobacterium and genetic transfer occur from the proteobacterium to the host , “the host providedthe membrane proteins for the import of substrates and enzymes for glycolysis, and ATP productionbegan under anaerobic conditions” (Paracer and Ahmadjian 83). As time progressed, the archaean hostbegan to utilize more complex and organic molecules instead of hydrogen and carbon dioxide.Evidences supporting the Endosymbiotic TheoryIn the following sections, information that point towards the SET will be discussed, and will be dividedbetween the two major eukaryotic organelles that are often considered- mitochondria and chloroplasts.
8MitochondriaFigure 4- A 3D cross-sectional view of a mitochondrion, which contains its own circular mitochondrial DNA,ribosomes (note that they are 70S), ATP synthase and double membrane. It is of no surprise that some speculate itsorigin from a prokaryotic cell, which also possesses a circular DNA molecule and ribosomes of the same sizeOne of the main claims that mitochondria might have originated from an ancestral proteobacterium isthat a mitochondrion possesses its own single, circular DNA molecule that is not associated withhistones, proteins that participate in the organization and wrapping of DNA in eukaryotic cells. Inhuman mitochondria, it consists of 37 genes (Endosymbiosis and the Origin of DomainEukaryotafossilmuseum.net)“16,569 base pairs of DNA in a closed circle which encodes 2 rRNAmolecules, 22 tRNA molecules and 13 polypeptides, with these polypeptides constructing proteincomplexes such as NADH dehydrogenase complex, ATP synthase, cytochrome c oxidase andcytochrome b in the inner mitochondrial membrane” (Endosymbiosis and The Origin of Eukaryotes
9users.rcn.com). This indeed bears an extremely close resemblance to a prokaryote, which also has itsown circular DNA that is free of histones. However, mitochondrial DNA in different organism do notshare the same size, with “two or three times more genome in plant mitochondria than in animalmitochondria” and “much of the plant mitochondrial DNA consists of noncoding sequences known asintrons” (Paracer and Ahmadjian 85). As prokaryotic DNA is not consisted of introns, despite thestriking similarity between the molecular features of mitochondria and prokaryotes, one cannot insistthat mitochondria definitely derived from ancestral eubacteria. By studying the mitochondria of fungus,it was found that the tRNA sequences are not similar to either prokaryotes or most eukaryotes (Paracerand Ahmadjian 85). Also, mitochondrial mRNA “have intervening sequences, or introns, along withpost-transcriptional processing” (Paracer and Ahmadjian 85) which is again not a feature spotted inprokaryotes but only in eukaryotes. However, one can still argue in favor of the SET that mitochondrialgenome consists of introns because of the transfer of genes between the host and the involvedendosymbiont, which results in changes in the genetic makeup of the proto-mitochondrion.Another piece of evidence pertaining to this matter is that mitochondria have 70S ribosomes that arealso found in prokaryotes but not in eukaryotes, and they have their own unique protein-synthesizingmechanism that again relates to prokaryotic cells. Antibiotics such as streptomycin block proteinsynthesis in bacteria but not in eukaryotic cells, and they too block protein synthesis in mitochondria;similarly, rifampicin inhibits both the RNA polymerase of bacteria and mitochondria but not that ofeukaryotes (Endosymbiosis and The Origin of Eukaryotes users.rcn.com). On the other hand, inhibitors
10such as diphtheria toxin act on protein synthesis of eukaryotes but not that of bacteria or mitochondriaEndosymbiosis and The Origin of Eukaryotes users.rcn.com). The fact that the translation machinery inmitochondria bears such likeness to that found in prokaryotes does help to validate the SET, but notfully justify it as it is not flawless. Pertaining to this evidence it is possible to counter it with theautogenous theory, which argues that mitochondria resemble prokaryotes because of mosaic evolution,where components within the compartments formed by invaginations of an ancestral prokaryotic cell’splasma membrane evolved much slower than the other parts.Figure 5- Electron microscope images of Rhodopseudomonas (left) and R. prowazekii (right); Rhodopseudomonasis a gram-negative bacteria that can display four modes of metabolism: photoheterotrophic, photoautotrophic,chemoheterotrophic and chemoautotrophic; R. prowazekii is also a gram-negative, intracellular (obligate) andaerobic alpha proteobacteria that is the causative agent of typhus; they are both genetically and molecularly relatedto mitochondriaPerhaps the most solid evidence that supports the SET, “the sequences of cytochromes, ferredoxin, and5S rRNA in mitochondria are similar to those of photosynthetic prokaryotes such asRhodopseudomonas” and by “sequencing rRNA from mitochondria of fungi, mice and humans, from
11nuclei of animals, yeast, and corn chloroplasts, and from E.coli that the greatest similarity in conservedsequences was between mitochondria and prokaryotes” (Paracer and Ahmadjian 85). Molecularsequencing also displays that proteobacteria are the closest modern day relatives to mitochondria, as“sequence data suggest that all extant mitochondria are derived from an ancestor of R. prowazekii(gram-negative proteobacteria) as the result of a single endosymbiotic event (2% of the protein-codinggenes inE.coli are found in all examined mitochondria).” (Berg, Tymoczko and Stryer 546)Another interesting evidence to note is that mitochondria is involved in the intrinsic apoptosis pathway,which is “initiated by the formation of the cytosolic apoptosome composed of Apaf-1 (protease),procaspase 9, and the cytochrome c released from mitochondria” (Jeong and Seol, “The role ofmitochondria in apoptosis”) in response to apoptotic signals. This reflects the interdependencedisplayed between the host and endosymbiont in ancient times after endosymbiosis had taken place forbillions of years, as a result of gene transfer between the two.
12ChloroplastsFigure 6- A two dimensional cross-section view of a chloroplast, which, similar to mitochondria, also possesses itssingle circular loop of DNA, 70S ribosomes and double membrane; however, they are unique in the sense that theyhave these extensive system of membranous sacs called thylakoids that actively engage in light-dependentreactions by containing photosynthetic pigmentsChloroplasts, like mitochondria, also have their own unique single, circular DNA. There are 128 genesthat include “duplicate genes encoding each of the four subunits of the rRNA, 37 genes encoding tRNA,4 genes encoding parts of the subunits of RNA polymerase, one gene encoding the large subunit of theenzyme RUBISCO (small subunit encoded by nuclear genome), 9 genes for components ofphotosystems I and II, 6 genes for parts of the ATP synthase and the rest of the genes for 19 of the 60proteins used to construct ribosome” (Endosymbiosis and The Origin of Eukaryotes users.rcn.com).Like mitochondria, this bears a resemblance to prokaryotic cells. Furthermore, the fact that chloroplastsrely on nuclear genes to encode their structural proteins (for instance the small subunit of the enzymeRUBISCO used in the Calvin Cycle) translated in the cytosol, which are then transported into the
13chloroplasts supports the interdependence that had resulted from billions of years of endosymbiosiswhere gene transfer took place and stripped the endosymbiont of the ability to live on its own. “Plastids(major double membrane organelles in plants that specialize in the storage and assembly of compoundsrequired by cells) typically contain some 60-100 genes, compared to cyanobacteria that have some1500 genes.” (Endosymbiosis and the Origin of Domain Eukaryotafossilmuseum.net) This againdemonstrates the gleaming possibility of the endosymbiont’s gene transfer to its host over the course ofendosymbiosis.Another evidence which supports chloroplasts originated as cyanobacteria that were engulfed byprokaryotic host cell capable of phagocytosis is that chloroplasts have double phospholipid bilayers.“The inner membrane represents the plasma membrane of the original prokaryotic symbiont, and theouter membrane either represents the vacuolar membrane formed by the host around the symbiont or isthe remnant outer membrane of the Gram-negative cyanobacterium that was the endosymbiont.”(Paracer and Ahmadjian 85) Other plastids such as chromoplasts and leucoplasts could have beenderived from secondary endosymbiosis, where heterotrophic eukaryotes proceeded to engulf otherphotosynthetic eukaryotes. For instance, “a group of unicellular, motile algae called cryptomonadsappear to be the evolutionary outcome of a nonphotosynthetic eukaryotic flagellate engulfing a red algaby endocytosis.” (Endosymbiosis and The Origin of Eukaryotes users.rcn.com); perhaps morespecifically, Paramecium bursaria, a unicellular eukaryote, engulfs green algae named zoochlorellaand develops a symbiotic relationship with it instead of digesting it. Zoochlorella provides
14P.bursariawith the ability to photosynthesize sugar, while they “extract nourishment from the hostwhen it is well fed and when they are deprived of light” and are “situated within individual vacuolesand these alga-vacuole complexes grow and divide at a rate compatible with that of the paramecium.”(Karakashian, “Symbiosis in Paramecium. Bursaria”) These are solid living examples ofendosymbiosis that provide insights as to the rise of eukaryotic cells. However, it is still possible toargue that plastids like chloroplasts arise as a result of secondary and even further invaginations of theancestral prokaryotic cell’s plasma membrane, resulting in the phenomenon that the “chloroplasts ofEuglena have three outer membranes, and those of brown algae and diatoms are surrounded by fourmembranes” (Paracer and Ahmadjian 85).Chloroplasts are very similar to mitochondria in the sense that they also possess 70S ribosomes foundcommonly in prokaryotes, and their own protein-synthesizing mechanism. For instance, Methionine isalways the first amino acid to be translated along the mRNA transcript; antibiotics such as streptomycinprevents protein synthesis in chloroplasts just as it is in bacteria and inhibitors such as diphtheria toxinhas no effect on protein synthesis unlike eukaryotes (Endosymbiosis and The Origin of Eukaryotesusers.rcn.com). Just as it was mentioned under the “mitochondria” section, one can argue that this doesnot support the SET by applying the autogenous theory, which simply considers the prokaryoticmolecular features of mitochondria and chloroplasts as a result of mosaic evolution.
15Perhaps the strongest piece of evidence for the SET regarding chloroplasts is from data collected frommolecular sequencing, such as “DNA-RNA hybridization of chloroplasts, nucleocytoplasm, andcyanobacteria and sequencing of plastocyanin, ferredoxin, and cytochrome c….have indicated a closerelationship between chloroplasts and blue-green prokaryotes.” (Paracer and Ahmadjian 86) Moreimportantly, the 5S and 16S sequences of chloroplast rRNA is much similar to that of cyanobacteriathan that of eukaryotes (Paracer and Ahmadjian 86).Conclusion and Discussion- Is the Serial Endosymbiotic Theory entirely credible?The information and evidences presented regarding the SET are largely overwhelming due to thebreadth of this topic, and the sections above barely provided a glimpse of arguments supporting andagainst this evolutionary theory. The fact that both mitochondria and chloroplasts greatly resembleprokaryotes both morphologically and molecularly, as they both possess double phospholipid bilayers,circular DNA molecule, 70S ribosomes and DNA/RNA sequences similar to prokaryotes, increases thecredibility of the endosymbiotic theory. These organelles are potentially the descendants of engulfedproteobacteria and cyanobacteria that goes back to billions of years ago. The sequencing of proteinsand nucleic acids had substantially proven the genetic link of these organelles to their ancestral cells;also, there are living organisms such as Paramecium bursaria that can be presented as living proof ofthe possibility of endosymbiosis as a way of mutually benefitting both the host and the endosymbiont.
16However, the SET is not without its flaws: “All evolutionary theories must offer an explanation inmechanistic terms of how it should or could have happened in order to be tested.” (Albert de Roos,“Acritique on the endosymbiotic theory for the origin of mitochondria”) The exact mechanisms of howthe genetic transfer from the endosymbiont to the host occurred, how organelles like mitochondria andchloroplasts acquired their double phospholipid membranes, how the interdepence between the twoevolved are not explained in this vaguely defined theory. The fact that endosymbiotic theory claimsmitochondria derived from the endocytosis of aerobic proteobacteria by anaerobic archaebacteriapresents a problem, as the host cell clearly could not tolerate any oxygen whilst the endosymbiontrequire oxygen to carry out aerobic respiration. There would also be a competition for energy resourcesas the endosymbiont requires ATP and would not easily transport them to the host cell. There is also theproblem that “the extensive gene transfer that is needed in the endosymbiotic theory would wreakhavoc in a complex genome since frequent insertion of random pieces of endosymbiont’s DNA woulddisrupt existing functions…the endosymbiotic theory is in contrast with the concept of gradualism thatforms the basis of modern evolutionary theory.” (Albert de Roos,“A critique on the endosymbiotictheory for the origin of mitochondria”) Pertaining to the evidence that mitochondria have circular DNAsimilar to that of bacteria, linear mitochondrial DNA molecules exist with “eukaryotic telomeres”.(Albert de Roos,“A critique on the endosymbiotic theory for the origin of mitochondria”) Althoughgenetic sequencing supports that mitochondria have a monophyletic origin from proteobacteria, “recentstudies of protists…indicate that the mitochondrion arose in a common ancestor of all extanteukaryotes and raise the possibility that this organelle originated at essentially the same time as the
17nuclear component of the eukaryotic cell rather than in a separate, subsequent event.” (Gray, Burgerand Lang, “Mitochondrial evolution”) In other words, the autogenous theory seems to explain the riseof mitochondria better as it claims that organelles form from invaginations of an ancestral prokaryoticcell’s plasma membrane. Speaking of the autogenous theory, it also appears to be more favorable as theevolution of eukaryotic organelles could be “driven by the advantages to sequester metabolic activity inspecialized compartments.” (Albert de Roos,“A critique on the endosymbiotic theory for the origin ofmitochondria”) The specific roles of the different organelles could have been based on the existingmetabolic functionalities, which were expanded and amplified gradually over time. In an addition, “themitochondrial genes could be derived from transposable elements, plastids or viruses and could comefrom either the nuclear genome or a bacterial genome.” (Albert de Roos,“A critique on theendosymbiotic theory for the origin of mitochondria”)In short, the true evolutionary origins of eukaryotic organelles is still undetermined due to the vastamount of time scientists are covering and the lack of substantial proof as to how every bit of themechanism worked, with no way to experimentally testify. Although the SET offers a probable theoryof endosymbiosis coupled by evolution that is backed by modern molecular data and observation, thereare still a lot of questions unanswered and missing links undiscovered. Other viable options such asautogenous theory present a simpler and more direct suggestion as to the rise of eukaryotic organelles,and while they are not without faults these alternative theories should be considered before the finalanswer is sealed.
18BibliographyAhmadjian, Vernon, and SurindarParacer.Symbiosis: An Introduction to Biological Associations.Hanover [N.H.: Published for Clark University by UP of New England, 1986. Print"Endosymbiosis and the Origin of Domain Eukaryota."Endosymbiosis. The Virtual Fossil Museum, 14Jan. 2012. Web. 01 Apr. 2013. <http://www.fossilmuseum.net/Evolution/Endosymbiosis.htm>."Endosymbiosis and The Origin of Eukaryotes." Endosymbiosis and The Origin of Eukaryotes. N.p., 31Jan. 2001. Web. 01 Apr. 2013.<http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/E/Endosymbiosis.html>.Gray, MW, Burger, G, and Lang, BF. "Mitochondrial Evolution."National Center for BiotechnologyInformation. U.S. National Library of Medicine, 05 Mar. 1999. Web. 01 Apr. 2013.<http://www.ncbi.nlm.nih.gov/pubmed/10066161?dopt=Abstract>.Jeong, Seon-Yong, and Dai-Wu Seol.The Role of Mitochondria in Apoptosis. Rep. BMB, n.d. Web. 01Apr. 2013. <http://bmbreports.org/jbmb/jbmb_files/%5B41-1%5D0801281812_11.pdf>.
19Karakashian, MW. "Symbiosis in Paramecium Bursaria."National Center for BiotechnologyInformation. U.S. National Library of Medicine, 31 Jan. 2002. Web. 01 Apr. 2013.<http://www.ncbi.nlm.nih.gov/pubmed/785659>.Roos, Albert De. "A Critique on the Endosymbiotic Theory for the Origin of Mitochondria - TelicThoughts."A Critique on the Endosymbiotic Theory for the Origin of Mitochondria - Telic Thoughts.Telic Thoughts, 17 Oct. 2007. Web. 01 Apr. 2013. <http://telicthoughts.com/a-critique-on-the-endosymbiotic-theory-for-the-origin-of-mitochondria/>.Stryer, Lubert, John L. Tymoczko, and Jeremy M. Berg.Biochemistry. New York: W.H. Freeman, 1988."The Endosymbiotic Theory."Endosymbiotic Theory. N.p., 14 Jan. 2002. Web. 01 Apr. 2013.<http://www.biology.iupui.edu/biocourses/N100/2k2endosymb.html>.