What exactly did Lynn Margulis contribute to science?

Lynn Margulis (wiki)

In my previous blogpost I remarked that Lynn Margulis deserved a Nobel prize for her endosymbiosis theory. This triggered a comment essentially denying the scientific merits of Lynn Margulis for evolutionary biology. It was claimed that Margulis was not the first to propose a symbiosis theory for the origin of mitochondria and chloroplasts and that she did not provide the DNA evidence. Here I provide evidence that during the critical 1960s and 70s mainstream science rejected and ridiculed the endosymbiosis theory and cytoplasmic heredity, whereas Margulis was defending this. Finally, I discovered two publications proving that she did publish DNA evidence.

Priorities, predecessors and prejudices

Indeed, there were predecessors of the symbiosis theory. However, previous publications were not generally known or, when they were noticed at all, they were ridiculed. For example, American biologist Ivan Wallin published in 1927 the book 'Symbionticism and the Origin of Species' proposing his version of the symbiosis theory. Unfortunately, "The reviews were so scathing that Wallin withdrew from scientific discourse. (...) To mainstream biologists, Wallin was little better than a crank." [1]. And according to wikipedia "Wallin and his works were largely forgotten, until Lynn Margulis introduced a more complete endosymbiotic theory with better evidences in 1967." [12].

Further, the Russian biologist Merezhkovsky "proposed that the organelles [chloroplasts] are actually cyanobacteria that took up residence in an early ancestor of plant cells. (...) That scenario was ignored or flatly rejected." (...) "But in 1966 decisive evidence for the validity of his proposal about mitochondria came with the discovery that these organelles have their own DNA." [2].

Additional evidence: the website 'Understanding Evolution' states: "Biologist Lynn Margulis first made the case for endosymbiosis in the 1960s, but for many years other biologists were skeptical. (...) Why should we think that a mitochondrion used to be a free-living organism in its own right? (...) Based on decades of accumulated evidence, the scientific community supports Margulis’ ideas: endosymbiosis is the best explanation for the evolution of the eukaryotic cell." [3].

Continuing from the same website:

"When one of her [Margulis] professors saw DNA inside chloroplasts, Margulis was not surprised. After all, that’s just what you’d expect from a symbiotic partner. Margulis spent much of the rest of the 1960s honing her argument that symbiosis was an unrecognized but major force in the evolution of cells. In 1970 she published her argument in the book The Origin of Eukaryotic Cells." [4].

Three big bangs that made genetics

Next, we need to place Margulis in the context of the history of genetics. In 1900 the rediscovery of Mendel made the first big bang in the biological sciences. The second was no less important: the identification of chromosomes as the physical location of Mendel's hypothetical factors. At the time this was called the Boveri–Sutton chromosome theory or the chromosome theory of inheritance. Nobody knows those name anymore. And it isn't a theory anymore. But it is an essential and necessary step in the construction of genetics as a science because the theory 1) located the hypothetical Mendelian factors on chromosomes, 2) showed elegantly that Mendels pairs of factors corresponded with pairs of chromosomes (homologous chromosomes). Amazingly, Mendel's theory predicted that we are all diploid creatures! Mendel never knew. The third big bang happened a decade before Margulis was trained as a biologist (1960s). That was the publication of the chemical structure of DNA by Watson and Crick in 1953. With a big bang molecular genetics became the most successful theory in the biological sciences. Together these 3 big bangs made genetics the most successful discipline within the biological sciences. Don't underestimate the feeling of triumph.

What is the relevance of all this? The relevance of these three discoveries for my argument is that chromosomes are located in the nucleus of the cell. In other words: 'genetics' was synonymous with chromosomal based heredity or nuclear heredity. This was the status of biology in the 1960s. Margulis was working in the shadow of mainstream nuclear genetics. She stubbornly decided to study extra-chromosomal inheritance or 'cytoplasmic heredity'. It is called 'cytoplasmic heredity' because mitochondria and chloroplasts are located in the cytoplasma of the cell. It was opposed to mainstream genetics in no less than 3 ways: non-chromosomal, non-nuclear (outside the nucleus or extra-nuclear) and non-Mendelian heredity. Cytoplasmic heredity was considered unimportant or controversial by the majority of geneticists. For example, Margulis quotes the Drosophila geneticist and Nobel Prize winner Thomas Hunt Morgan: 

"From the point of view of heredity, the cytoplasm of a cell can safely be ignored." [5] 

Margulis commented: "I considered it an arrogant oversimplification." I agree. In general it is 'amusing' and instructive to browse through old genetics textbooks to get a feeling of the thinking at the time [14]. Everybody knew that heredity was exclusively located in the nucleus of eukaryotes. It was an established truth. It wasn't even called 'nuclear genetics' or 'nuclear genes', because everybody knew that the hereditary material was located in the nucleus! 

Illustrative for the status of science is the statement of John Jinks (1964) in his monograph Extrachromosomal inheritance: 

"There have been increasingly persistent claims that DNA is present in structures such as plastids and mitochondria." [10]. 

 Please note that 'claims' means not proven ideas.

In A History of Genetics the genetics pioneer A.H. Sturtevant (1965) wrote: 

Sturtevant 1965
A History of Genetics

"Quite recently it has been found that there is DNA in plastids and at least some mitochondria. ... It may therefore be supposed that these bodies carry genes of the same nature as those in the chromosomes" [6]. 

It must have been a shocking discovery for those unaware of the symbiosis theory. See for example what historian of science Jan Sapp wrote:

"The symbiotic theory of eukaryotic organelles attracted criticism from many sides during the 1970s. Crucial evidence was lacking. ... Leading cell biologists also argued that theories about eukaryotic cell origins were unscientific because they could not be proven." (!) [7]. (my emphasis)

That was how biologists used to think at the time. In 1972 Tribe and Whittaker wrote in their monograph Chloroplasts and Mitochondria:

"Up until the last few years, it was generally assumed that the total information necessary for the synthesis of a cell resided in the DNA of the nucleus." [8].

Indeed! Remarkably, at the time it was uncertain whether mitochondria and chloroplasts could divide at all, or whether they were synthesized de novo (from scratch). It was also a problem how much of the DNA in mitochondria and chloroplasts was functional at all and, if functional, what role it played. Furthermore, the authors point out two major objections to the symbiotic theory [9]. So, my point is that one should not ignore the status of biology at the time Margulis was working.

DNA evidence!

The blog commenter also claimed that Margulis did not provide DNA evidence for the symbiosis theory. This is not correct. I discovered an ignored publication: 

L. Sagan (1961) Evidence for cytoplasmic DNA in Euglena gracillis [11] in the  Journal of Protozoology. 

This publication is not listed in the Lynn Margulis wikipedia page. Unfortunately, google scholar returns the title only, not the contents, not even an abstract. However, the title is revealing [12]. The most reliable and exemplary demonstration of DNA in plastids is given by S. Nass and M.K. Nass (1963) [13]. From the fact that they perform extensive control experiments, it is clear that they are aware that extraordinary claims require extraordinary evidence. They wanted to exclude any contamination with nuclear DNA or other false positives. At the time these authors were not interested in evolutionary explanations and did not interpret the DNA as evidence for symbiosis theory. They were busy unraveling the structure and function of cellular organelles. No time for speculation. This shows that the discovery of DNA in mitochondria occurred independent of the symbiosis theory. On the other hand knowledge of the symbiosis theory would be an excellent reason to search for DNA in mitochondria. Nass & Nass came incredibly close to the discovery of an evolutionary symbiosis theory with this quote : "a structure homologous with the mitochondrion is the entire bacterial cell." That statement is still not a symbiosis theory. The most far-reaching conclusion they dared to draw was:

"In any case, the presence of D N A in mitochondria appears to require modification and extension of some generally accepted hypotheses of cell function which consider the nucleus to be the exclusive site of cellular D N A and genetic information." [13].

Indeed. Clearly, the authors doubted a well established truth. It was the first step towards a symbiosis theory. Lynn Margulis devoted herself to developing a comprehensive symbiosis theory.

Principles of Population Genetics

'Symbiosis' is still a sensitive subject. A stunning demonstration is the textbook Principles of Population Genetics [15]. Many pages are devoted to describing all genetics aspects of mitochondrial DNA (mtDNA) and chloroplast DNA (cpDNA). And those aspects are without doubt useful for understanding certain aspects of evolution. However, (endo)symbiosis is never mentioned. It is not that mitochondria are absent. It is not that mitochondrial DNA is absent. No. It is the word 'Symbiosis' that is absent. It doesn't occur in the index. I find that really puzzling. Is symbiosis so far beyond the scope of population genetics? It seems symbiosis does not fit in the theory of population genetics. Could it be that the origin of mitochondria and chloroplasts can not even be described by the most advanced mathematical population genetic models? If true, how relevant is population genetics for understanding all the facts of evolution of life on earth?

A second paper of DNA evidence by Lynn Margulis is:

L Sagan, Y Ben-Shaul, HT Epstein, JA Schiff (1965) 'Studies of chloroplast development in Euglena. XI. Radioautographic localization of chloroplast DNA', Plant physiology 1965 Nov;40(6):1257–1260. (pdf). (Note: at the time Lynn Margulis published under the name L. Sagan because she was married with Carl Sagan.)

This publication does not occur in wikipedia either. The authors conclude that chloroplasts (like mitochondria) contain DNA and RNA; they are self-duplicating bodies which do not arise de novo; the DNA represents a multigenic hereditary system which is not derived from the nucleus and is, in part, responsible for the biochemical properties of the organelle. In her 1967 paper On The Origin of Mitosing Cells she made the prediction that all eukaryotes should contain mitochondrial DNA and all eukaryotic plants should contain chloroplast DNA. Also in that 1967 paper she refers to  (Sagan, L., 1964, unpublished data) [16].

Conclusion

The claim that Margulis was not the first to propose a symbiosis theory is right. However, that is not the point. The point is that the symbiosis theory was disregarded at the time when Margulis started working on the subject. The evidence presented here shows that it was risky and unpopular to study and publish about the symbiosis theory during the 1960s and 1970s. Margulis (1967,1970) elaborated the theory and provided enough initial evidence to justify further research into the theory. Furthermore, I showed that the claim that she did not come up with DNA evidence is wrong. The symbiosis theory provided a correct evolutionary explanation for why there is DNA in mitochondria and chloroplasts.

Now we have reached this conclusion, it is useful to compare Darwin and Margulis. Darwin did not prove his theory in 1859. Neither was he the first to propose a theory of evolution. And his theory was very controversial at the time. But, Darwin provided enough reasons for his contemporaries to take the theory serious. Subsequent generations of biologists have shown that the theory was indeed worth investigating and came with abundant evidence. It is now an accepted theory. The same holds for the endosymbiosis theory.

I am not the only one with a positive assessment of the contribution of Lynn Margulis to the theory of evolution. For example Michael Ruse and Joseph Travis [17] wrote:

"But it was Margulis who took up the idea, now known as the endosymbiotic theory, offered microscopic evidence, and pushed it (in her book Origin of Eukaryotic Cells, 1970) until it became orthodoxy.".

Notes

1. Lynn Margulis, Dorion Sagan (1997) Slanted Truths chapter 4, page 50.  
2. Lynn Margulis, Dorion Sagan (1997) Slanted Truths, chapter 3, page 38.
3. Website Understanding Evolution page 'Evidence for endosymbiosis' of the Berkeley museum of paleontology. (by the way, very good website!)
4. The full title: Origin of eukaryotic cells;: Evidence and research implications for a theory of the origin and evolution of microbial, plant, and animal cells on the Precambrian earth (1970) Yale University Press. (still available at amazon). See also: the website Understanding Evolution page Endosymbiosis: Lynn Margulis
5. Lynn Margulis (1998) The Symbiotic Planet. A new look at evolution, hardback, page 22.
6. A.H. Sturtevant (1965) A History of Genetics page 125 chapter 19 'Maternal Effects'. Sturtevant refers to: Gibor and Granick (1964) Science 145: 890-897. This publication is also referred to by Margulis (1967). 
7. Jan Sapp (2003) Genesis. The Evolution of Biology. page 246.
8. Michael Tribe, Peter Whittaker (1972) Chloroplasts and Mitochondria. page 54.
9. idem page 57.
10. John L. Jinks (1964) Extrachromosomal inheritance. Prentice-Hall. p.89. Dutch: John L. Jinks (1968) Extrachromosomale erfelijkheid. Het Spectrum. paperback, pagina 132. He did not refer to publications. 
11. Sagan, L, Evidence for cytoplasmic DNA in Euglena gracillis, Journal of Protozoology 8 4: 20 (1961). Euglena is a single-celled eukaryote. 
12. I found the publication via the references in Lynn Sagan (1967) On the origin of mitosing cells (her first publication. It is a large, complex, wide-ranging publication). She writes: "A plethora of recent studies elegantly reviewed by Gibor & Granick": A. Gibor and S. Granick (1964) Plastids and Mitochondria: Inheritable Systems: Do plastids and mitochondria contain a chromosome which controls their multiplication and development? Science 28 Aug 1964. Only the Abstract is available, but, amazingly, I discovered in the References of G&G: "L. Sagan (1961)" [11].  
13. S. Nass and M.K. Nass (1963) Intramitochondrial fibers with DNA characteristics. II. Enzymatic and Other Hydrolytic Treatments. J Cell Biol 1963 Dec 1;19(3):613–629. A pdf is available.
14. For example the genetics textbook Srb, Owen, Edgar (1965) General Genetics, Second edition, hardback, which I still have on my bookshelf, contains a chapter 'Extrachromosomal and Epigenetic Systems'. That is great. But browsing through the pages reveals that the most important criterion for extrachromosomal inheritance is simply that it does not obey the laws of Mendel and must be located in the cytoplasma. Mitochondria and chloroplasts are mentioned, but at the time it was impossible to prove that these organelles contain DNA. It was an open question whether extrachromosomal heredity relies on 'coded information'. "Extrachromosomal heredity has reality, and manifests itself in almost bewildering variety." (p.345). There is no mention of '(endo)symbiosis'. How amusing and useful a little time travel can be!
15. Hartl & Clark (1997) Principles of Population Genetics, third edition. Sinauer Associates. hardback. 
16. "This is quite analogous to the presence of two distinct DNA bands of nearly equal size found in CsCl density gradient runs on DNA isolated from Paramecium bursaria (Sagan, L., 1964, unpublished data). 
17. Michael Ruse, Joseph Travis (2009) Evolution. The First Four Billion Years,  Harvard University Press, hardback, 979 pages. page 712-713. This is just one example. The influential evolutionary biologist John Maynard Smith recognized her contribution to science; see my review of Lynn Margulis.