Let's look at a famous paper by Jack Lester King and Thomas Hughes Jukes.1 The title of the paper is "Non-Darwinian Evolution" and it was published 44 years ago in the May 16, 1969 issue of Science [read it at: Science 164:788-798].
The subtitle of the paper is "Most evolutionary change in proteins may be due to neutral mutations and genetic drift" but that's not what I want to talk about. This paper is among the first to predict the presence of large amounts of junk DNA in our genome. King and Jukes didn't call it "junk"—that term was introduced by Susumu Ohno in 1972—but that doesn't matter. When King and Jukes talk about "superfluous DNA" they mean "junk."
Here's the relevant part of the paper ...
Different proteins evolve at different rates, and different sites within specific proteins evolve at different rates. It is possible that these differences reflect differential mutability of the DNA itself, but to us this seems unlikely. It is more likely that proteins, and sites within proteins, differ with regard to the stringency of their requirements. The average rate of evolutionary change as shown in Table 1 is 16 × 10-10 substitution per codon per species per year.Note that 44 years ago scientists had a pretty good understanding of mutation rate and they realized that many amino acid substitutions would be neutral. Nevertheless, some mutations are lethal and if the entire human genome consists of genes (coding) then the lethal mutation rate will be too high for survival. This is the genetic load argument.
Kimura (4) has estimated, in agreement with Jukes (37), that total molecular evolution in vertebrate species proceeds at the rate of about one amino acid substitution every 2 years. Arguing that Darwinian evolution at that rate would require greater selection pressure than any species can afford, Kimura concluded that most amino acid changes must be due to the passive fixation of selectively neutral mutations.
While we tend to agree with this conclusion, there are several reasons for questioning the arguments on which it was based. Kimura's estimate was deliberately conservative in some respects. The estimate was based (i) on comparisons of the beta chains of horse and human hemoglobins, which appear to have about an average rate of evolutionary change; (ii) on studies of cytochrome c, which is a relatively slowly evolving protein; and (iii) on a minimum estimate based on unsequenced analysis of triosephosphate dehydrogenase, this probably being a gross underestimate of the true evolutionary rate for that enzyme. The average rate of evolution per codon in the completely sequenced proteins listed in Table 1 is five times Kimura's conservative underestimate. If the rate per codon is extrapolated to the entire haploid DNA genome of 4 × 109 nucleotide pairs, as has been done previously (4, 37), it would appear that mammalian evolution is proceeding at the rate of about two allele substitutions per year. In relatively long-lived mammals this may be 20 substitutions per species per generation; in the human species, this is an evolutionary rate of nearly 60 amino acid substitutions per generation, implying a genome mutation rate including 60 neutral amino acid substitutions per gamete. For several reasons this seems much too high.
For one thing, about 4 percent of base substitutions result in chain-terminating codons; 60 amino acid substitutions imply about three chain-terminating mutations per gamete. Most chain-terminating mutations, if they occur in structural genes, are lethal, or at least produce nonfunctional alleles which have to be eliminated through natural selection. No organism having three lethal or severely deleterious mutations per gamete can survive. In addition, frame-shift mutations, also lethal in structural genes, appear to occur about as frequently as chain-terminating mutations (30), and certainly some of the amino acid substitutions are lethal or biologically harmful. Indeed, as we attempt to demonstrate below, it is unlikely that more than about 10 percent of all mutations are selectively neutral.
A second error is the assumption that all or most mammalian DNA consists of structural genes. Older estimates (see 38) of maximum gene number in mammals rarely exceed 40,000 genes per haploid genome. If the average gene consists of 1000 nucleotide pairs, extrapolation from the estimated evolutionary rate of 16 × 10-10 substitution per codon per year gives one amino acid substitution per species per 50 years. This is a far more believable figure. But only 4 × 107 nucleotide pairs, or 1 percent of the mammalian genome, is thus accounted for. Either 99 percent of mammalian DNA is not true genetic material, in the sense that it is not capable of transmitting mutational changes which affect the phenotype, or 40,000 genes is a gross underestimate of the total gene number.This begins the discussion about whether most of our genome is functional. The data seems to suggest that a large percentage of the human genome is immune to mutational changes.
Rates of spontaneous mutation to recessive lethal and visible mutants in mammals are of the order of 10-6 to 10-5 per locus per generation (38). If there are 40,000 genes, the total rate of mutation to lethal or nonfunctional alleles would be between 4 and 40 percent per gamete. From this consideration alone, it is clear that there cannot be many more than 40,000 genes.
In extensive studies of the spontaneous mutation rate of Drosophila melanogaster, the average lethal mutation rate was 3 × 10-6 per locus and 10-2 per genome (39). Thus, the fruit fly has about 3000 loci that are capable of mutating to lethal alleles. If only a third of all loci are capable of mutating to lethal alleles under laboratory conditions, there may be perhaps 10,000 Drosophila cistrons. If the average cistron size is 1000 nucleotides, this accounts for about 10 percent of Drosophila DNA (8), since drosophilas have much less DNA per cell than mammals have.It's important to note that back in 1969, scientists had pretty good ideas about genome sizes in different species. It's also important to understand that scientists like King and Jukes were perfectly capable of combining information from a number of different species in order to reach a general conclusion that would be valuable for all species. That form of argument is not as widely practiced today.
There is more direct evidence for the existence of nongenetic DNA. Heterochromatin is known to be nearly devoid of specific genetic information, yet it accounts for about a third of the DNA of those species in which it is cytologically detectable. About 30 percent of mammalian DNA consists of highly repetitive sequences of unknown function (9). In some species there are varying numbers of supernumerary chromosomes that appear to be of no survival value to the organism.Even 44 years ago, scientists knew about repetitive DNA and other genome sequences that were noncoding. This should dispel the notion that informed scientists ever ignorant about the composition of noncoding DNA.
Perhaps the most compelling argument for the existence of superfluous DNA is the wide range in the DNA content of vertebrate cells (40, 41). The average mammalian cell contains more than twice the DNA of the chicken cell and almost four times that of the cell of the gar pike. The cell of the bullfrog contains twice as much DNA as that of the toad, and two and a half times as much as that of a man, while the cell of a lungfish has a DNA content 17 times that of the human cell and almost 60 times that of the pike cell. Can it be that these wide divergences in DNA content reflect wide divergences in the number of functional genes? This hardly seems likely.King and Jukes make three arguments for superfluous (junk) DNA: (1) genetic load, (2) known examples of repetitive, noncoding DNA, and (3) the C-value paradox. These are examples of positive arguments for the existence of substantial quantities of junk DNA in our genome. It should put to rest any claims that the existence of junk DNA was based entirely on ignorance.
King and Jukes were aware of counter-arguments. The most important of these was the claim that a substantial percentage of the genome is transcribed. Remember, this is 44 years ago.
On the other hand, a substantial proportion of mammalian DNA is capable of forming hybrids with specific messenger RNA in vitro (42). Possibly, as Callan suggests (40), numerous nonheritable copies of the essential genetic material are created anew each generation. These multiple copies would transmit specific information by way of messenger RNA, but would not be true genetic material in that they would not transmit information to future generations and would not be directly involved in evolutionary processes. Another important possibility is that much of mammalian DNA is involved in the complexities of the immune response (26).As is the case today, the fact that more than 1% of the genome is transcribed was not going to falsify the idea that most of the human genome does not carry genetic information.
Keep in mind that the title of this paper is "Non-Darwinian Evolution." Neither King nor Jukes thought of themselves as strict "Darwinists." The idea of junk (superfluous) DNA was promoted by scientists who understood neutral mutations and random genetic drift. Then, as now, strict Darwinists did not like the idea that most of our genome is junk.
1. Jukes earned his Ph.D. in biochemistry in my department in 1933.