Vol 13, Issue 5, 838-844, May 2003
LETTER
Neutral Substitutions Occur at a Faster Rate in Exons Than in Noncoding DNA in Primate Genomes
Sankar Subramanian and
Sudhir Kumar1
Center for Evolutionary Functional Genomics, Arizona Biodesign
Institute and Department of Biology, Arizona State University,
Tempe, Arizona 85287-1501, USA
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ABSTRACT
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Point mutation rates in exons (synonymous sites) and noncoding
(introns and intergenic) regions are generally assumed to be the same.
However, comparative sequence analyses of synonymous substitutions in
exons (81 genes) and that of long intergenic fragments (141.3 kbp) of
human and chimpanzee genomes reveal a 30%–60% higher mutation rate
in exons than in noncoding DNA. We propose a differential CpG content
hypothesis to explain this fundamental, and seemingly unintuitive,
pattern. We find that the increased exonic rate is the result of the
relative overabundance of synonymous sites involved in CpG
dinucleotides, as the evolutionary divergence in non-CpG sites is
similar in noncoding DNA and synonymous sites of exons. Expectations
and predictions of our hypothesis are confirmed in comparisons
involving more distantly related species, including human–orangutan,
human–baboon, and human–macaque. Our results suggest an underlying
mechanism for higher mutation rate in GC-rich genomic regions, predict
nonlinear accumulation of mutations in pseudogenes over time, and
provide a possible explanation for the observed higher diversity of
single nucleotide polymorphisms (SNPs) in the synonymous sites of exons
compared to the noncoding regions.
Point mutation is a major source of genetic
variation and evolution. The rate at which these mutations arrive in a
genome is routinely estimated in mutation accumulation experiments and
in disease mutation analysis (Drake et al. 1998 ; Giannelli et al. 1999;
Lynch et al. 1999; Denver et al. 2000). In comparative sequence
analysis, mutation rates are inferred by estimating the rate at which
neutral substitutions accumulate, because the rate of mutation and
substitution is expected to be the same for neutral mutations (Kimura
1983; Kondrashov and Crow 1993; Nei and Kumar 2000). Pseudogenes,
introns, intergenic regions, and synonymous sites in exons are largely
free from selection in mammals and are therefore routinely used in
comparative sequence analysis for this purpose (Wolfe et al. 1989; Li
and Graur 1991; Keightley and Eyre-Walker 2000; Nachman and Crowell
2000; Zhao et al. 2000; Kumar and Subramanian 2002).
A survey of mutation rate estimates obtained from the synonymous sites
of protein coding genes and noncoding DNA for the human and chimpanzee
comparison reveal a significant rate difference in these two types of
genomic regions. For instance, we find that the analyses of pseudogenes
(Li and Tanimura 1987; Nachman and Crowell 2000; Martinez-Arias et al.
2001) and introns (Bergstrom et al. 1999; Chen et al. 2001) have
produced much lower estimates of evolutionary divergence than those
observed in synonymous sites in protein coding genes (Li and Tanimura
1987; Wolfe et al. 1989; Easteal 1991; Keightley and Eyre-Walker 2000;
Duret et al. 2002; Kumar and Subramanian 2002). Also, intergenic DNA
(excluding regulatory sites), which is expected to mutate at a rate
similar to that for the coding DNA, also consistently shows much
smaller evolutionary divergences than those obtained from coding
sequences (Bohossian et al. 2000; Zhao et al. 2000; Chen et al. 2001;
Mathews et al. 2001; Yu et al. 2001). This observation is not a random
chance occurrence due to paucity of data, because these studies have
involved direct comparisons of long intergenic regions (Bohossian et
al. 2000; Zhao et al. 2000; Chen et al. 2001) and relatively large
numbers of coding genes (Wolfe et al. 1989; Keightley and Eyre-Walker
2000; Duret et al. 2002; Kumar and Subramanian 2002). This difference
between coding and noncoding regions in interspecies comparisons is
also seen in within-species analysis of single nucleotide polymorphisms
(SNPs) that show higher nucleotide diversity in the synonymous sites of
exons compared to the noncoding DNA (Moriyama and Powell 1996; Cargill
et al. 1999; Halushka et al. 1999; Zwick et al. 2000).
In order to systematically examine the extent and nature of mutation
rate differences that have potentially given rise to the observed
differences between synonymous sites of exons and noncoding DNA, we
present an analysis of an extensive data set containing long stretches
of intergenic DNA, introns, and pseudogenes as well as a large number
of coding genes from human and other primates.
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RESULTS
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Patterns of Mutation Rate Difference
We estimated neutral evolutionary distances between human and
chimpanzee genomes using the 141.3 kbp of intergenic DNA, 19
pseudogenes, 81 protein-coding genes, and introns from 48 of these
protein-coding genes (Table 1; Methods).
The synonymous divergence was significantly higher than the sequence
divergence observed in different noncoding genomic regions
(P < 0.002; Table 1). Similar patterns of difference are
seen in the human–orangutan and human–baboon comparisons, suggesting
that this phenomenon is common to hominids and old world monkeys (Table
1). It is unlikely that this difference can be explained by increased
purifying selection in noncoding regions, as the data set analyzed
contains rather long intergenic blocks (10–60 kb) from different human
chromosomes and a large number of introns and pseudogenes. All of these
vastly different genomic regions are unlikely to be under strong
purifying selection with a similar intensity.
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Table 1. Relative Rates of Evolutionary Divergence Between Synonymous Sites in
Coding DNA and Different Types of Noncoding
Regions
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The Differential CpG Content Hypothesis
We explored an alternative hypothesis to explain the observed
difference in mutation rates, which is based on the known phenomenon of
hypermutability of methylated CpG dinucleotides to TpG (Coulondre et
al. 1978; Bird 1980; Britten et al. 1988; Ponger et al. 2001). As
formulated, the differential-CpG-content hypothesis states that the
difference in the CpG content is responsible for the observed mutation
rate difference between exons and noncoding DNA. To test this
hypothesis, we re-estimated the evolutionary divergence in exons,
pseudogenes, introns, and intergenic sequences by excluding synonymous
sites involved in the CpG dinucleotides in each case. The mutation rate
difference among coding and noncoding DNA disappeared, as the ratio of
synonymous distance to that observed in pseudogenes, introns, or
intergenic sequences is now close to 1 (Table 1). This result is
further confirmed in analyses of 48 genes from human–chimpanzee
comparison and 25 genes from human–baboon comparison for which the
exon and intron sequences were available for the same genes (Table 1).
The differential contribution of CpG mutations to the overall mutation
rate in noncoding DNA and the synonymous sites of exons is evident from
Figure 1. The observed difference in
evolutionary divergence correlates with the larger proportion of CpG
dinucleotides in exons than those in the noncoding DNA (cf. open bars
in Fig. 1A,B). This result holds true even when we normalize the CpG
content by accounting for GC-content bias (data not shown). The finding
that divergence at non-CpG sites (black bars in Fig. 1A) is almost the
same in the synonymous sites of exons and in different types of
noncoding regions indicates that the difference in CpG content, rather
than selection, is largely responsible for the observed difference in
the neutral evolutionary divergence in exons and noncoding DNA.
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Figure 1. (A) Evolutionary divergence per 100 sites and (B) CpG
contents of synonymous sites in exons, pseudogenes, introns, and
intergenic DNA for the human–chimpanzee comparison. The mean and the
standard error estimates shown were obtained from 81 protein coding
genes, 19 pseudogenes, introns of 48 genes, and four intergenic blocks.
In (A), the total height of each column (including black and
open bars) corresponds to the overall evolutionary divergence. The
black portion in each column depicts divergence with CpG sites
excluded, and therefore, the open bars show the contribution of
mutations at CpG sites to the overall divergence. In (B), the
CpG content is the proportion of sites involved in the CpG dinucleotide
configuration. For noncoding DNA, all sites were included for the
estimation of CpG content and evolutionary divergence, whereas for
exons only the fourfold-degenerate sites were included (see Methods).
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Directionality of the CpG Mutations
The depletion of CpG dinucleotides in vertebrate genomes has been
attributed to the directional hypermutability of these dinucleotides to
TpG or CpA (Bird 1980; Cooper and Krawczak 1989; Karlin and Mrazek
1997; Hendrich et al. 1999). We examined this directionality of the CpG
mutations (Bird 1980) in an analysis of introns and synonymous sites of
22 genes from human and chimpanzee. For these genes, baboon sequences
were used to identify the direction of the change at the CpG sites in
the human lineage and in the chimpanzee lineage. Results show that for
a total of 4600 sites that were potentially CpG in the ancestral
sequence (based on the most parsimonious reconstructions), 5.72%
experienced CpG  TpG/CpA changes. In contrast, only 0.82% of
ancestrally TpG or CpA show a change to CpG. This roughly estimated
 sevenfold difference in forward and reverse mutation rates is in
agreement with previous studies on disease mutations and comparative
sequence analysis, which have shown CpG mutations to be 5–20 times
more frequent than other types of point mutations (Batzer et al. 1990;
Yang et al. 1996; Krawczak et al. 1998; Giannelli et al. 1999;
Ebersberger et al. 2002).
Pseudogenes As the Missing Link Between Coding and Noncoding DNA
We can also track the evolutionary dynamics of CpG depletion by
comparing pseudogenes with their functional counterparts (e.g., Sved
and Bird 1990). This provides a glimpse into the process of decay of
CpG dinucleotides, as recently emerged pseudogenes are expected to show
much higher CpG content than the older pseudogenes. An analysis of 39
pseudogenes from the human genome indeed shows that the overall
evolutionary divergence correlates negatively with the ratio of CpG
content between the pseudogenes and their functional counterparts
(R2 = –0.52, P < 0.01; Fig. 2A). This
clear pattern of CpG decay over time is further demonstrated in the
analysis of the pseudogenes created at different times from the same
functional genes (Fig. 2B,C).
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Figure 2. Relationship between evolutionary divergence and the ratio of CpG
contents in pseudogenes and their functional counterparts. (A)
Analysis of 39 human pseudogenes [R2 = 0.52;
P < 0.01], (B) seven pseudogenes created from the
same Arginino succinate synthetase gene
[R2 = 0.90; P < 0.01], and
(C) five pseudogenes of the same Beta tubulin Q gene
[R2 = 0.73; P < 0.05]. The relative
CpG content is the ratio of the overall CpG content in the pseudogene
divided by overall CpG content of the functional gene.
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A comparison of Figure 1A and B also shows that the evolutionary
divergence as well as the CpG content of pseudogenes are at an average
intermediate compared to those at synonymous sites and noncoding DNA.
This result is now explained by the finding that the pseudogenes
created recently from the functional coding genes have higher CpG
content than introns and intergenic regions (Fig. 2). However, as
pseudogenes become older their CpG content declines until it reaches
the level present in the surrounding noncoding DNA. This also means
that the average rate of mutation per site in a pseudogene will
decrease with time, because the number of CpG dinucleotides will
decrease over time. Therefore, there are no linear mutation clocks for
pseudogene evolution.
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DISCUSSION
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It is now clear that there is a significant difference in the
evolutionary divergence between the noncoding and the synonymous sites
of the coding DNA. This difference correlates with differences in the
content of the hypermutable CpG dinucleotides in these two types of
genomic DNA. This is a causal relationship, as the removal of CpG
dinucleotides from data eliminates the difference. The higher CpG
content in codon positions is already well appreciated as the observed
versus expected (O/E) ratio of CpG contents in the 1st–2nd, 2nd–3rd,
and 3rd–1st positions is known to be higher than that for the genomic
sequences (Sved and Bird 1990; Caccio et al. 1997; Jabbari and Bernardi
1998; Halushka et al. 1999). In the data set of introns and exons of 48
genes from human and chimpanzee comparison, we find that the noncoding
DNA shows an O/E ratio of 0.23, whereas the synonymous sites in exons
show an O/E ratio of 0.40. This is an almost twofold difference and is
likely caused by the fact that one of the nucleotides of the CpG
dinucleotides involving a fourfold-degenerate site is under purifying
selection, as they are always from the 1st or 2nd codon position. This
increases the probability of occurrence and maintenance of CpG
dinucleotides in exons compared to noncoding DNA, where both
nucleotides in the CpG dinucleotides are free to undergo neutral
nucleotide substitution. This results in a higher average mutational
input per synonymous site, as synonymous sites are largely free to
evolve irrespective of the selection on the adjacent codon positions.
The knowledge of CpG content-mediated difference in mutation rates also
provides new insights into reasons behind a number of previous
observations. For instance, a significant positive correlation has been
reported between the GC content of the 3rd codon position and the
synonymous substitution in primates (Bielawski et al. 2000). This is
often thought to be due to differences in rates of mutations among
regions with differing base compositions (Smith et al. 2002; Yi et al.
2002). However, GC-rich regions are expected to contain more CpG
dinucleotides than GC-poor regions. It is therefore possible that
differences in CpG content in GC-poor and GC-rich regions are a major
factor in the observed synonymous distance difference. This is indeed
the case, as revealed in an analysis of 93 genes from our
human–macaque comparison (Fig. 3A,B). The
positive correlation between GC content and the synonymous distance
becomes rather small when synonymous sites involved in CpG
dinucleotides are excluded. These results are consistent with the
reports of CpG-mediated correlation between the heterozygosity and GC
content of the human SNPs (Sachidanandam et al. 2001).
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Figure 3. (A) Relationship between evolutionary divergence and GC
content, GC4, at synonymous sites of 93 genes from the
human–macaque comparison; R2 = 0.125,
P < 0.01. (B) Relationship between evolutionary
divergence and GC4 after excluding fourfold-degenerate sites
involved in CpG dinucleotides; R2 = 0.01,
P > 0.05.
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Furthermore, there have been reports of positive correlation between
evolutionary distances observed at synonymous sites and the noncoding
DNA for the human–mouse genome comparison (Hardison et al. 2003). This
positive correlation (R2 = 0.48,
P < 0.01) is also reflected in our analysis of 25 genes of
human and baboon (Fig. 4A). However after
the exclusion of CpG sites, this correlation reduces significantly
(Fig. 4B; R2 = 0.12), suggesting that the CpG
mutations mediate this relationship in these primate genomes. In fact,
the R2 value for this relationship drops down to
0.04 if we remove just a single outlier (Fig. 4B). Thus, the actual
rates of mutation at non-CpG sites, which comprise >95% of noncoding
DNA and >90% of synonymous sites, are quite similar in GC-rich and
GC-poor regions. This result is in contrast to some of the previous
studies of human–mouse genome comparisons (e.g., Waterston et al.
2002; Hardison et al. 2003), which presumably removed the influence of
CpG mutations when estimating neutral distances. However, when the time
of species divergence is very large, as is the case for human and mouse
(90–110 million years ago, see Kumar and Hedges 1998; Hedges and Kumar
2003; Springer et al. 2003), the identification of CpG sites as well as
the assessment of CpG mutations on overall divergence becomes
unreliable (see also Methods). Therefore, it is important to use
closely related species to examine the mutation rate variation across
the genome.
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Figure 4. (A) Relationship between synonymous divergence in exons and
evolutionary divergence in introns; R2 = 0.511,
P < 0.01. (B) Relationship between synonymous
divergence in exons and evolutionary divergence in introns after
removing CpG sites; R2 = 0.137,
P > 0.05. With the rightmost outlier excluded,
R2 = 0.06, P > 0.05.
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Finally, the demonstration of significant differences in evolutionary
rates in exonic and noncoding DNA due to CpG-content difference
provides a potential explanation for the observed 30%–40% higher
diversity in the human SNPs in the fourfold-degenerate sites of exons
compared to noncoding DNA (Cargill et al. 1999; Halushka et al. 1999).
This difference is similar to that observed in the interspecies
comparisons (Table 1). It was also reflected in an analysis of 6109
human SNPs, which we mapped to fourfold-degenerate sites in the
reference human sequence (August 7, 2002 release of dbSNP from NCBI was
used). We found that 39.5% of all SNPs at the fourfold-degenerate
sites were of CpG  TpG/CpA types, which is close to the proportion
of variable sites with CpG TpG/CpA difference in the interspecies
comparisons (39.0%–48.4%). Therefore, although the
synonymous sites of exons and noncoding DNA may be used for estimating
coalescence times of population polymorphisms (and alleles), they will
require the use of different mutation rates unless the CpG mutations
are excluded.
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METHODS
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Data Mining and Assembly
Protein coding genes used in this study were obtained from the
HOVERGEN database, release 36 (Duret et al. 1994) and from the GenBank.
Phylogenetic trees of 8627 HOVERGEN gene families were constructed from
amino acid sequence alignments using the neighbor-joining (NJ) method
in MEGA2 (Kumar et al. 2001). The cDNA sequence alignments for
orthologous sequence sets were then generated using amino acid sequence
alignments as guides. NJ trees were scanned automatically, followed by
manual inspection, to identify orthologous sequence sets. We enforced
strict orthology definitions by considering sequences to be orthologous
only if no gene duplication events were detected since their divergence
from the most recent common ancestor. Gene duplication events were
identified by comparing the gene family tree with the species tree from
NCBI (see also Zmasek and Eddy 2001). All gene families containing
fewer than three sequences were excluded. This produced a data set
consisting of 33, 38, 93, and six orthologous pairs of sequences for
human–chimpanzee, human–orangutan, human–macaque, and human–baboon
comparisons, respectively. We also obtained a set of 19 autosomal
pseudogenes through searching the literature (Nachman and Crowell 2000;
Chen et al. 2001). In addition, orthologous noncoding intergenic
segments belonging to human, chimpanzee, and orangutan genomes were
obtained (Bohossian et al. 2000; Zhao et al. 2000). To compare the
evolutionary divergence at the neutral sites of exons and introns of
the same gene, we obtained (from GenBank) 48 genes for the
human–chimpanzee comparison and 25 genes for the human–baboon
comparison. When not available, the intron–exon boundaries of
chimpanzee and baboon genes were identified by using the corresponding
human exons in the BLASTZ software (Schwartz et al. 2003). For
intergenic sequence comparisons, five large contigs (human: AC002066,
AC002080, chimpanzee: AC087253, AC087512, baboon: AC084730) were
obtained from GenBank. All repeat elements in these contigs were
identified using RepeatMasker
(http://ftp.genome.washington.edu/cgi-bin/RepeatMasker) and removed
prior to the sequence alignment. CpG islands in the upstream regions
are usually unmethylated and are not hypermutable (Bird 1999).
Therefore, they were also excluded from further analyses. (However, our
results remained the same whether or not they were excluded). Also, the
coding genes known from the annotated human genome were removed, along
with the 5-kb flanking regions immediately upstream and downstream of
each gene to exclude potential regulatory regions. This resulted in
intergenic regions with sizes of 52.6 kbp (AC002066 and AC087512) and
65.0 kbp (AC002080 and AC087253) for the human–chimpanzee comparison,
and 38.0 kbp (AC002066 and AC084730) for the human–baboon comparison.
All noncoding sequences were aligned using BLASTZ.
For analysis of CpG depletion in pseudogenes, we used 19 pseudogenes
for the human–chimpanzee comparison. In addition, we obtained 116
human pseudogenes from the GenBank. Using local BLAST with 31,289 human
cDNA (obtained from ftp://ftp.ncbi.nih.gov/genomes/H_sapiens/), we
found the functional counterparts of these pseudogenes, which had an
e-value of 0. We restricted our analysis to the pseudo-functional gene
pairs for which evolutionary distance was small (d < 0.2).
Our final data set contains 39 pseudogene-functional gene pairs.
Estimation of Evolutionary Divergence
Evolutionary divergences were computed using only the
fourfold-degenerate sites for coding sequences and all sites for
pseudogenes, introns, and intergenic regions. We took a stringent
approach in identifying fourfold-degenerate sites by selecting only
those sites that have remained fourfold-degenerate in both the species
compared. A genomic synonymous distance estimate was obtained by
concatenating all coding genes for a given species pair. Evolutionary
distances were estimated for pseudogenes and introns by concatenating
all the respective sequences. Correction for multiple hits was done
using the Tamura-Nei method, which accounts for transition/transversion
and base composition biases (Tamura and Nei 1993). CpG content was
estimated as the number of sites involved in the CpG configuration
divided by the total number of sites. For exons, only the
fourfold-degenerate sites involved in the CpG configuration (G of the
CpG dinucleotide in the second and third codon positions and C of the
CpG dinucleotide in the third and first [adjacent] codon positions)
were used. For noncoding DNA (including pseudogenes and introns) and
the coding genes used for the CpG depletion study (Fig. 2), all sites
involved in CpG dinucleotides were included. In the
pseudogene-functional gene comparisons, we excluded all CpG sites when
estimating the evolutionary divergence, in order to avoid introducing
spurious negative correlation.
Note that the existing methods to correct for multiple hits in
evolutionary distances estimation do not account for the CpG
substitutions, which occur at a much higher rate than other mutations
and with a different pattern. Use of Tamura-Nei method (Tamura and Nei
1993) is expected to lead to underestimation in this case. However in
our analysis we have used only closely related species. In such cases
the choice of a simpler substitution model is known to produce minimal
underestimation (see Nei and Kumar 2000). For the current scenario, we
examined the extent of underestimation by conducting a computer
simulation in which two identical sequences with a prespecified CpG
content were made to evolve over time, and the evolutionary divergence
between sequences was estimated using the Tamura-Nei distance. CpG
substitutions were made to occur at 10 times the rate of all other
substitutions, and the total actual number of substitutions was
recorded. The estimated numbers of substitutions obtained using
Tamura-Nei (1993) distance were compared to the actual number of
substitutions at difference levels of sequence divergence for the
sequence length of 10,000. Simulations were conducted with different
GC contents (25%, 50%, and 75%). We found that for
sequence divergences considered here (<0.1 per site), the Tamura-Nei
distances were less than 10% underestimates of the true distance (in
each case they were slight underestimates; but they were large for
divergences >0.1 per site). This means that the difference in rates
between exons and other genomic regions reported here are conservative
estimates, as the sequence divergence from exons with higher CpG
content will be underestimated more than that for the CpG-poor
noncoding regions.
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WEB SITE REFERENCES
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http://ftp.genome.washington.edu/cgi-bin/RepeatMasker;
RepeatMasker Web server.
http://www.nisc.nih.gov; NIH intramural sequencing center.
ftp://ftp.ncbi.nih.gov/genomes/H_sapiens/; Human genomic sequences at
National Center for Biotechnology Information.
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Acknowledgements
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We thank Philip Hedrick, Koichiro Tamura, Michael Rosenberg, Robert
Friedman, Alan Filipski, Rekha Iyer, and Patrick Kolb for helpful
discussions and comments. We also thank Alan Filipski for help with
computer simulations, Michael Rosenberg for analysis of SNP data,
Balaji Ramanujam for some data retrieval, and David Cutler, Phil Green,
and an anonymous reviewer for insightful comments. Many of the genomic
contigs used in this study were generated by the NIH Intramural
Sequencing Center (http://www.nisc.nih.gov). This research was
supported by research grants from the NIH, NSF, and the
Burroughs-Wellcome Fund to S.K.
The publication costs of
this article were defrayed in part by payment of page charges. This
article must therefore be hereby marked "advertisement" in
accordance with 18 USC section 1734 solely to indicate this fact.
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Footnotes
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1 Corresponding author. 
E-MAIL s.kumar{at}asu.edu; FAX(480) 965-2519.
Article and publication are at
http://www.genome.org/cgi/doi/10.1101/gr.1152803.
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REFERENCES
|
|---|
Batzer, M.A., Kilroy, G.E., Richard, P.E., Shaikh, T.H., Desselle, T.D., Hoppens, C.L., and Deininger, P.L. 1990. Structure and variability of recently inserted Alu family members. Nucleic Acids Res. 18: 6793-6798.[Abstract/Free Full Text]
Bergstrom, T.F., Erlandsson, R., Engkvist, H., Josefsson, A., Erlich, H.A., and Gyllensten, U. 1999. Phylogenetic history of hominoid DRB loci and alleles inferred from intron sequences. Immunol. Rev. 167: 351-365.[CrossRef][Medline]
Bielawski, J.P., Dunn, K.A., and Yang, Z. 2000. Rates of nucleotide substitution and mammalian nuclear gene evolution. Approximate and maximum-likelihood methods lead to different conclusions. Genetics 156: 1299-1308.[Abstract/Free Full Text]
Bird, A. 1999. DNA methylation de novo. Science 286: 2287-2288.[Free Full Text]
Bird, A.P. 1980. DNA methylation and the frequency of CpG in animal DNA. Nucleic Acids Res. 8: 1499-1504.[Abstract/Free Full Text]
Bohossian, H.B., Skaletsky, H., and Page, D.C. 2000. Unexpectedly similar rates of nucleotide substitution found in male and female hominids. Nature 406: 622-625.[CrossRef][Medline]
Britten, R.J., Baron, W.F., Stout, D.B., and Davidson, E.H. 1988. Sources and evolution of human Alu repeated sequences. Proc. Natl. Acad. Sci. 85: 4770-4774.[Abstract/Free Full Text]
Caccio, S., Jabbari, K., Matassi, G., Guermonprez, F., Desgres, J., and Bernardi, G. 1997. Methylation patterns in the isochores of vertebrate genomes. Gene 205: 119-124.[CrossRef][Medline]
Cargill, M., Altshuler, D., Ireland, J., Sklar, P., Ardlie, K., Patil, N., Shaw, N., Lane, C.R., Lim, E.P., Kalyanaraman, N., et al. 1999. Characterization of single-nucleotide polymorphisms in coding regions of human genes. Nat. Genet. 22: 231-238.[CrossRef][Medline]
Chen, F.C., Vallender, E.J., Wang, H., Tzeng, C.S., and Li, W.H. 2001. Genomic divergence between human and chimpanzee estimated from large-scale alignments of genomic sequences. J. Hered. 92: 481-489.[Abstract/Free Full Text]
Cooper, D.N. and Krawczak, M. 1989. Cytosine methylation and the fate of CpG dinucleotides in vertebrate genomes. Hum. Genet. 83: 181-188.[CrossRef][Medline]
Coulondre, C., Miller, J.H., Farabaugh, P.J., and Gilbert, W. 1978. Molecular basis of base substitution hotspots in Escherichia coli. Nature 274: 775-780.[CrossRef][Medline]
Denver, D.R., Morris, K., Lynch, M., Vassilieva, L.L., and Thomas, W.K. 2000. High direct estimate of the mutation rate in the mitochondrial genome of Caenorhabditis elegans. Science 289: 2342-2344.[Abstract/Free Full Text]
Drake, J.W., Charlesworth, B., Charlesworth, D., and Crow, J.F. 1998. Rates of spontaneous mutation. Genetics 148: 1667-1686.[Abstract/Free Full Text]
Duret, L., Mouchiroud, D., and Gouy, M. 1994. HOVERGEN: A database of homologous vertebrate genes. Nucleic Acids Res. 22: 2360-2365.[Abstract/Free Full Text]
Duret, L., Semon, M., Piganeau, G., Mouchiroud, D., and Galtier, N. 2002. Vanishing GC-rich isochores in mammalian genomes. Genetics 162: 1837-1847.[Abstract/Free Full Text]
Easteal, S. 1991. The relative rate of cDNA evolution in primates. Mol. Biol. Evol. 8: 115-127.[Abstract]
Ebersberger, I., Metzler, D., Schwarz, C., and Paabo, S. 2002. Genome-wide comparison of DNA sequences between humans and chimpanzees. Am. J. Hum. Genet. 70: 1490-1497.[CrossRef][Medline]
Giannelli, F., Anagnostopoulos, T., and Green, P.M. 1999. Mutation rates in humans. II. Sporadic mutation-specific rates and rate of detrimental human mutations inferred from hemophilia B. Am. J. Hum. Genet. 65: 1580-1587.[CrossRef][Medline]
Halushka, M.K., Fan, J.B., Bentley, K., Hsie, L., Shen, N., Weder, A., Cooper, R., Lipshutz, R., and Chakravarti, A. 1999. Patterns of single-nucleotide polymorphisms in candidate genes for blood-pressure homeostasis. Nat. Genet. 22: 239-247.[CrossRef][Medline]
Hardison, R.C., Roskin, K.M., Yang, S., Diekhans, M., Kent, W.J., Weber, R., Elnitski, L., Li, J., O'Connor, M., Kolbe, D., et al. 2003. Covariation in frequencies of substitution, deletion, transposition and recombination during eutherian evolution. Genome Res. 13: 13-26.[Abstract/Free Full Text]
Hedges, S.B. and Kumar, S. 2003. Genomic clocks and evolutionary timescales. Trends Genet. (in press).
Hendrich, B., Hardeland, U., Ng, H.H., Jiricny, J., and Bird, A. 1999. The thymine glycosylase MBD4 can bind to the product of deamination at methylated CpG sites. Nature 401: 301-304.[CrossRef][Medline]
Jabbari, K. and Bernardi, G. 1998. CpG doublets, CpG islands and Alu repeats in long human DNA sequences from different isochore families. Gene 224: 123-127.[CrossRef][Medline]
Karlin, S. and Mrazek, J. 1997. Compositional differences within and between eukaryotic genomes. Proc. Natl. Acad. Sci. 94: 10227-10232.[Abstract/Free Full Text]
Keightley, P.D. and Eyre-Walker, A. 2000. Deleterious mutations and the evolution of sex. Science 290: 331-333.[Abstract/Free Full Text]
Kimura, M., 1983. The neutral theory of molecular evolution. Cambridge University Press, Cambridge, UK.
Kondrashov, A.S. and Crow, J.F. 1993. A molecular approach to estimating the human deleterious mutation rate. Hum. Mutat. 2: 229-234.[CrossRef][Medline]
Krawczak, M., Ball, E.V., and Cooper, D.N. 1998. Neighboring-nucleotide effects on the rates of germ-line single-base-pair substitution in human genes. Am. J. Hum. Genet. 63: 474-488.[CrossRef][Medline]
Kumar, S. and Hedges, B. 1998. A molecular timescale for vertebrate evolution. Nature 392: 917-919.
Kumar, S. and Subramanian, S. 2002. Mutation rates in mammalian genomes. Proc. Natl. Acad. Sci. 99: 803-808.[Abstract/Free Full Text]
Kumar, S., Tamura, K., Jakobsen, I.B., and Nei, M. 2001. MEGA2: Molecular evolutionary genetics analysis software. Bioinformatics 17: 1-2.[Free Full Text]
Li, W.-H. and Graur, D., 1991. Fundamentals of molecular evolution. Sinauer Associates, Sunderland, MA.
Li, W.-H. and Tanimura, M. 1987. The molecular clock runs more slowly in man than in apes and monkeys. Nature 326: 93-96.[CrossRef][Medline]
Lynch, M., Blanchard, J., Houle, D., Kibota, T., Schultz, S., Vassilieva, L., and Willis, J. 1999. Perspective: Spontaneous deleterious mutation. Evolution 53: 645-663.[CrossRef]
Martinez-Arias, R., Calafell, F., Mateu, E., Comas, D., Andres, A., and Bertranpetit, J. 2001. Sequence variability of a human pseudogene. Genome Res. 11: 1071-1085.[Abstract/Free Full Text]
Mathews, D.J., Kashuk, C., Brightwell, G., Eichler, E.E., and Chakravarti, A. 2001. Sequence variation within the fragile X locus. Genome Res. 11: 1382-1391.[Abstract/Free Full Text]
Moriyama, E.N. and Powell, J.R. 1996. Intraspecific nuclear DNA variation in Drosophila. Mol. Biol. Evol. 13: 261-277.[Abstract]
Nachman, M.W. and Crowell, S.L. 2000. Estimate of the mutation rate per nucleotide in humans. Genetics 156: 297-304.[Abstract/Free Full Text]
Nei, M. and Kumar, S., 2000. Molecular evolution and phylogenetics. Oxford University Press, New York, NY.
Ponger, L., Duret, L., and Mouchiroud, D. 2001. Determinants of CpG islands: Expression in early embryo and isochore structure. Genome Res. 11: 1854-1860.[Abstract/Free Full Text]
Sachidanandam, R., Weissman, D., Schmidt, S.C., Kakol, J.M., Stein, L.D., Marth, G., Sherry, S., Mullikin, J.C., Mortimore, B.J., Willey, D.L., et al. 2001. A map of human genome sequence variation containing 1.42 million single nucleotide polymorphisms. Nature 409: 928-933.[CrossRef][Medline]
Schwartz, S., Kent, W.J., Smit, A., Zhang, Z., Baertsch, R., Hardison, R.C., Haussler, D., and Miller, W. 2003. Human–mouse alignments with BLASTZ. Genome Res. 13: 103-107.
Smith, N.G., Webster, M.T., and Ellegren, H. 2002. Deterministic mutation rate variation in the human genome. Genome Res. 12: 1350-1356.[Abstract/Free Full Text]
Springer, M.S., Murphy, W.J., Eizirik, E., and O'Brien, S.J. 2003. Placental mammal diversification and the Cretaceous-Tertiary boundary. Proc. Natl. Acad. Sci. 100: 1056-1061.[Abstract/Free Full Text]
Sved, J. and Bird, A. 1990. The expected equilibrium of the CpG dinucleotide in vertebrate genomes under a mutation model. Proc. Natl. Acad. Sci. 87: 4692-4696.[Abstract/Free Full Text]
Tamura, K. and Nei, M. 1993. Estimation of the number of nucleotide substitutions in the control region of mitochondrial DNA in humans and chimpanzees. Mol. Biol. Evol. 10: 512-526.[Abstract]
Waterston, R.H., Lindblad-Toh, K., Birney, E., Rogers, J., Abril, J.F., Agarwal, P., Agarwala, R., Ainscough, R., Alexandersson, M., An, P., et al. 2002. Initial sequencing and comparative analysis of the mouse genome. Nature 420: 520-562.[CrossRef][Medline]
Wolfe, K.H., Sharp, P.M., and Li, W.-H. 1989. Mutation rates differ among regions of the mammalian genome. Nature 337: 283-285.[CrossRef][Medline]
Yang, A.S., Gonzalgo, M.L., Zingg, J.M., Millar, R.P., Buckley, J.D., and Jones, P.A. 1996. The rate of CpG mutation in Alu repetitive elements within the p53 tumor suppressor gene in the primate germline. J. Mol. Biol. 258: 240-250.[CrossRef][Medline]
Yi, S., Ellsworth, D.L., and Li, W.H. 2002. Slow molecular clocks in old world monkeys, apes, and humans. Mol. Biol. Evol. 19: 2191-2198.[Abstract/Free Full Text]
Yu, N., Zhao, Z., Fu, Y.X., Sambuughin, N., Ramsay, M., Jenkins, T., Leskinen, E., Patthy, L., Jorde, L.B., Kuromori, T., et al. 2001. Global patterns of human DNA sequence variation in a 10-kb region on chromosome 1. Mol. Biol. Evol. 18: 214-222.[Abstract/Free Full Text]
Zhao, Z., Jin, L., Fu, Y.X., Ramsay, M., Jenkins, T., Leskinen, E., Pamilo, P., Trexler, M., Patthy, L., Jorde, L.B., et al. 2000. Worldwide DNA sequence variation in a 10-kilobase noncoding region on human chromosome 22. Proc. Natl. Acad. Sci. 97: 11354-11358.[Abstract/Free Full Text]
Zmasek, C.M. and Eddy, S.R. 2001. A simple algorithm to infer gene duplication and speciation events on a gene tree. Bioinformatics 17: 821-828.[Abstract/Free Full Text]
Zwick, M.E., Cutler, D.J., and Chakravarti, A. 2000. Patterns of genetic variation in Mendelian and complex traits. Annu. Rev. Genomics Hum. Genet. 1: 387-407.[CrossRef][Medline]
Received January 3, 2003;
accepted in revised format March 11, 2003.
13:838-844 © by 2003 Cold Spring Harbor Laboratory Press ISSN 1088-9051/03 $5.00
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