Lc

Patient D-x

Patient D-y

| DNA sequences from patients D and F are no more similar to that from the dentist than to those from local controls (LC). These patients were probably not infected by the dentist.

23.20 Evolutionary tree showing the relationships of HIV isolates from a dentist, seven of his patients (A through G), and other HIV-positive persons from the same region (local controls, LC). The letters x and y represent different isolates from the same patient. The phylogeny is based on DNA sequences taken from the envelope gene of the virus. Viral sequences from patients A, B, C, E, and G cluster with those of the dentist, indicating a close evolutionary relationship. Sequences from patients D and F, along with those of local controls, are more distantly related. [C. Ou et al. Molecular epidemiology of HIV transmission in a dental practice, Science 2 56(1992): 1167.]

determine the number of nucleotides that differ between the two sequences. We might compare the growth-hormone sequences for mice and rats, which diverged from a common ancestor some 15 million years ago. From the number of different nucleotides in their growth-hormone genes, we compute the number of nucleotide substitutions that must have taken place since they diverged. Because the same site may have mutated more than once, the number of nucleotide substitutions is larger than the number of nucleotide differences in two sequences; so special mathematical methods have been developed for inferring the actual number of substitutions likely to have taken place.

When we have the number of nucleotide substitutions per nucleotide site, we divide by the amount of evolutionary time that separates the two organisms (usually obtained from the fossil record) to obtain an overall rate of nucleotide substitution. For the mouse and rat growth-hormone gene, the overall rate of nucleotide substitution is approximately 8 X 10—9 substitutions per site per year.

Nucleotide changes in a gene that alter the amino acid sequence of a protein are referred to as nonsynonymous substitutions. Nucleotide changes, particularly those at the third position of the codon, that do not alter the amino acid sequence are called synonymous substitutions. The rate of nonsynonymous substitution varies widely among mammalian genes. The rate for the a-actin protein is only 0.01 X 10—9 substitutions per site per year, whereas the rate for interferon 7 is 2.79 X 10—9, about 1000 times as high. The rate of synonymous substitution also varies among genes, but not to the extent of variation in the nonsynony-mous rate. For most protein-encoding genes, the synonymous rate of change is considerably higher than the non-synonymous rate because synonymous mutations are tolerated by natural selection (Table 23.10). Nonsynony-mous mutations, on the other hand, alter the amino acid sequence of the protein and in many cases are detrimental to the fitness of the organism, so most of these mutations are eliminated by natural selection.

Different parts of a gene also evolve at different rates, with the highest rates of substitutions in regions of the gene that have the least effect on function, such as the third position of a codon, flanking regions, and introns (< Figure 23.21). The 5' and 3' flanking regions of genes are not transcribed into RNA, and therefore substitutions in these regions do not alter the amino acid sequence of the protein, although they may affect gene expression (see Chapter 16). Rates of substitution in introns are nearly as high. Although these nucleotides do not encode amino acids, introns must be spliced out of the pre-mRNA for a functional protein to be produced, and particular sequences are required at the 5' splice site, 3' splice site, and branch point for correct splicing (see Chapter 14).

Substitution rates are somewhat lower in the 5' and 3' untranslated regions of a gene. These regions are transcribed into RNA but do not encode amino acids. The 5' untranslated region contains the ribosome-binding site, which is essential for translation, and the 3' untranslated region contains sequences that may function in regulating mRNA stability and translation; so substitutions in these regions may have deleterious effects on organismal fitness and will not be tolerated.

Table 23.10 Rates of nonsynonymous and synonymous substitutions in mammalian genes based on human-rodent comparisons

Gene a-Actin p-Actin Albumin Aldolase A Apoprotein E Creatine kinase Erythropoietin a-Globin p-Globin

Growth hormone Histone 3

Immunoglobulin heavy chain (variable region)

Insulin

Interferon a1

Interferon 7

Luteinizing hormone

Somatostatin-28

Nonsynonymous Rate (per Site per 109 Years)

0 0

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