Wednesday, May 6, 2009

The molecular clock should never have been invented in the first place for macroevolution

Two kinds of sequence alignment can be made using the same set of sequence data.  The first aligns a recently evolved organism such as a mammal against those simpler or less complex species that evolved earlier such as amphibians and fishes.  The second aligns a simpler outgroup organism such as fishes against those more complex sister species that appeared later such as amphibians and mammals.  The first alignment indicates a near linear correlation between genetic distance and time of divergence, implying indirectly a constant mutation rate among different species.  The second alignment shows the genetic equidistance result where sister species are approximately equidistant to the simpler outgroup. This directly triggered the idea of constant mutation rate among different species.  Since both alignments use the same sequence data set, certain information may be revealed by either alone.  But the data that most directly and obviously support the interpretation of a constant mutation rate is the genetic equidistance result. 

The molecular clock hypothesis was first informally proposed by Zuckerkandl and Pauling in 1962 based largely on data from the first alignment [1].  Margoliash in 1963 performed both alignments and made a formal statement of the molecular clock after noticing the genetic equidistance result [2, 3].  “It appears that the number of residue differences between cytochrome c of any two species is mostly conditioned by the time elapsed since the lines of evolution leading to these two species originally diverged. If this is correct, the cytochrome c of all mammals should be equally different from the cytochrome c of all birds.  Since fish diverges from the main stem of vertebrate evolution earlier than ether birds or mammals, the cytochrome c of both mammals and birds should be equally different from the cytochrome c of fish.  Similarly, all vertebrate cytochrome c should be equally different from the yeast protein.”

The results of both alignments have two features.  One is obvious: distance in terms of percent identity, which directly provoked the clock idea.  The other is the overlap feature.  In the post of April 30th, 2009, I explained the overlap feature of the genetic equidistance result.  Here, I show that the first kind of alignment performed by Zuckerkandl and Pauling also shows the overlap feature, as would be expected since both alignments use the same sequence information and should tell the same story.  The clock idea should never have been invented in the first place if Zuckerkandl and Pauling had paid attention to this feature.


Again, I use cytochrome c of yeast (Sc), drosophila (Dm), and human (Hs) as an example.  What Zuckerkandl and Pauling had found, when applied in our cytochrome c case here, is that human is closer to drosophila than to yeast.  Human differs from drosophila in 22 positions and from yeast in 36 positions.  The overlap feature in this case is that most of 22 variant positions between human and drosophila are also variant between human and yeast.  This can be easily illustrated in the following alignment:




                 *..:** .:*  :* ****** ** **.******::**::*** *::** ** 





                *.: *.*:.: *** *********** * *:** ::* ***:***.**  

The result shows that 17 of the 22 are also variant between human and yeast (these 17 positions are colored in purple and green).  The fact that the overlap is not 100% is because residues conserved due to common adaptation to environment between human and drosophila are different from those between human and yeast. 

The molecular clock predicts: 

The chance for a position to be different between human and yeast is 36/102.

The chance for a position to be different between human and drosophila is 22/102.

The number of overlap positions: 36/102 x 22/102 x 102 = 7.76. far short of 17.

There are only 7 positions as underlined below that are absolutely conserved among bacteria, yeast, plants, nematodes, and human.



So, a most realistic calculation of overlap should be 36/95 x 22/95 x 95 = 8.3 residues, far short of 17.


Even if we generously grant that 40 residues are absolutely non-neutral or non-variable, we still only get 36/62 x 22/62 x 62 = 12.77, short of 17. 

Again, Zuckerkandl, Pauling, and Margoliash all could have noticed the overlap feature.  If they had done that 46 years ago, the molecular clock (vastly different species have very similar mutation rates) would never have been invented in the first place for macroevolution.  It may have been invented for studying microevolution (identical or very similar species have very similar mutation rates) and may still apply in some cases of microevolution.  But its impact on the understanding of molecular evolution would be trivial. 


I thank my college classmate Dr. Wei Shen for providing the alignment picture shown here, and for many valuable discussions. 



1.         Zuckerkandl E, Pauling L: Molecular disease, evolution, and genetic heterogeneity, Horizons in Biochemistry. New York: Academic Press; 1962.

2.         Margoliash E: Primary structure and evolution of cytochrome c. Proc Natl Acad Sci 1963, 50:672-679.

3.         Kumar S: Molecular clocks: four decades of evolution. Nat Rev Genet 2005, 6:654-662.

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