A molecular view of primate phylogeny and important systematic and evolutionary questions

Phylogenetic analysis of extensive nucleotide sequence data from primate beta-globin gene clusters elucidates the systematics and evolution of the order Primates and reveals that rates of accumulation of mutations vary by as much as a factor of seven among different primate lineages. The picture of primate phylogeny from DNA sequences clarifies many ambiguities of the morphological picture. In the molecular picture, dwarf and brown lemurs group together into superfamily Lemuroidea, Lemuroidea and Lorisoidea into suborder Strepsirhini, and Tarsius and Anthropoidea into suborder Haplorhini. The molecular picture also provides both significant evidence for a human-chimpanzee clade that narrowly excludes gorilla and overwhelming evidence for the gorilla-chimpanzee-human clade within Hominoidea. Rates of DNA sequence evolution appear to have been fastest in the early primates ancestral to Anthropoidea and next fastest on the lorisoid branch. Rates were slowest over the past 25 Myr of hominoid descent, suggesting that mechanisms lowering the mutation rate evolved in correlation with lengthened life spans.     

Evolutionary rate variation in Old World monkeys

We analysed over 8 million base pairs of bacterial artificial chromosome-based sequence alignments of four Old World monkeys and the human genome. Our findings are as follows. (i) Genomic divergences among several Old World monkeys mirror those between well-studied hominoids. (ii) The X-chromosome evolves slower than autosomes, in accord with ‘male-driven evolution’. However, the degree of male mutation bias is lower in Old World monkeys than in hominoids. (iii) Evolutionary rates vary significantly between lineages. The baboon branch shows a particularly slow molecular evolution. Thus, lineage-specific evolutionary rate variation is a common theme of primate genome evolution. (iv) In contrast to the overall pattern, mutations originating from DNA methylation exhibit little variation between lineages. Our study illustrates the potential of primates as a model system to investigate genome evolution, in particular to elucidate molecular mechanisms of substitution rate variation.     

Forty Million Years of Independent Evolution: A Mitochondrial Gene and Its Corresponding Nuclear Pseudogene in Primates

Sequences from nuclear mitochondrial pseudogenes (numts) that originated by transfer of genetic information from mitochondria to the nucleus offer a unique opportunity to compare different regimes of molecular evolution. Analyzing a 1621-nt-long numt of the rRNA specifying mitochondrial DNA residing on human chromosome 3 and its corresponding mitochondrial gene in 18 anthropoid primates, we were able to retrace about 40 MY of primate rDNA evolutionary history. The results illustrate strengths and weaknesses of mtDNA data sets in reconstructing and dating the phylogenetic history of primates. We were able to show the following. In contrast to numt-DNA, (1) the nucleotide composition of mtDNA changed dramatically in the different primate lineages. This is assumed to lead to significant misinterpretations of the mitochondrial evolutionary history. (2) Due to the nucleotide compositional plasticity of primate mtDNA, the phylogenetic reconstruction combining mitochondrial and nuclear sequences is unlikely to yield reliable information for either tree topologies or branch lengths. This is because a major part of the underlying sequence evolution model — the nucleotide composition — is undergoing dramatic change in different mitochondrial lineages. We propose that this problem is also expressed in the occasional unexpected long branches leading to the “common ancestor” of orthologous numt sequences of different primate taxa. (3) The heterogeneous and lineage-specific evolution of mitochondrial sequences in primates renders molecular dating based on primate mtDNA problematic, whereas the numt sequences provide a much more reliable base for dating.[Reviewing Editor: Dr. Rafael Zardoya]     

Molecular phylogeny of the family of apes and humans

The morphological picture of primate phylogeny has not unambiguously identified the nearest outgroup of Anthropoidea and has not resolved the branching pattern within Hominoidea. The molecular picture provides more resolution and clarifies the systematics of Hominoidea. Protein and DNA evidence divides Hominoidea into Hylobatidae (gibbons) and Hominidae, family Hominidae into Ponginae (orangutan) and Homininae, and subfamily Homininae into two tribes, one for Gorilla, and the other for Pan (chimpanzee) and Homo. Parsimony and maximum likelihood analyses, carried out on orthologous noncoding nucleotide sequences from primate beta-globin gene clusters, provide significant evidence for the human-chimpanzee tribe and overwhelming evidence for the human-chimpanzee-gorilla clade. These analyses also indicate that the rate of molecular evolution became slower in hominoids than in other primates and mammals.     

Evolution of a repeat sequence in the parathyroid hormone-related peptide gene in primates

A polymorphism of the variable number of tandem repeat (VNTR) type is located 97 bp downstream of exon VI of the parathyroid hormone-related peptide (PTHrP) gene in humans. The repeat unit has the general sequence G(TA)_nC, where n equals 4–11. In order to characterize the evolutionary history of this VNTR, we initially tested for its presence in 13 different species representing four main groups of living primates. The sequence is present in the human, great apes, and Old World monkeys, but not in New World monkeys; and this region failed to PCR amplify in the Loris group. Thus, the evolution of the sequence as part of the PTHrP gene started at least 25–35 millions years ago, after divergence of the Old World and New World monkeys, but before divergence of Old World monkeys and great apes and humans. The structural changes occurring during evolution are characterized by a relatively high degree of sequence divergence. In general, the tandem repeat region tends to be longer and more complex in higher primates with the repeat unit motifs all being based on a TA-dinucleotide repeat sequence. Intra-species variability of the locus was demonstrated only in humans and gorilla. The divergence of the TA-dinucleotide repeat sequence and the variable mutation rates observed in different primate species are in contrast to the relative conservation of the flanking sequences during primate evolution. This suggests that the nature of the TA-dinucleotide repeat sequence, rather than its flanking sequences, is responsible for generating variability. Particular features of the sequence may allow it to form stable secondary structures during DNA replication, and this, in turn, could promote slipped-strand mispairing to occur.     

The relative rate of DNA evolution in primates

In 73 relative-rate tests involving the sequences of 17 genes between humans and six nonhuman primate taxa, there is only one significant (P less than 0.01) difference in evolutionary rate--i.e., that between human and Old World-monkey psi eta-globin genes. No evolutionary rate difference between humans and Old World monkeys is evident from analysis of 18 other genes with a total length of 6 kb. This and the comparison, between humans and other primate taxa, of new extended psi eta-globin sequences suggest that earlier observations of evolutionary-rate differences between humans and other primates were based on differences that are peculiar to psi eta-globin and that are not representative of the whole genome, which appears to be evolving at a stochastically uniform rate. This is supported by whole-genome single-copy DNA and mitochondrial DNA comparisons, neither of which shows any evidence of evolutionary-rate variation among primate taxa. Uniformity in the evolutionary rate of the DNA of primate and other mammalian taxa is inconsistent with current mammalian fossil-record interpretation. Either there has been a general slowing down in rate across lineages or the fossil record has been misinterpreted.     

Ancestral Population Sizes and Species Divergence Times in the Primate Lineage on the Basis of Intron and BAC End Sequences

The effective sizes of ancestral populations and species divergence times of six primate species (humans, chimpanzees, gorillas, orangutans, and representatives of Old World monkeys and New World monkeys) are estimated by applying the two-species maximum likelihood (ML) method to intron sequences of 20 different loci. Examination of rate heterogeneity of nucleotide substitutions and intragenic recombination identifies five outrageous loci (ODC1, GHR, HBE, INS, and HBG). The estimated ancestral polymorphism ranges from 0.21 to 0.96% at major divergences in primate evolution. One exceptionally low polymorphism occurs when African and Asian apes diverged. However, taking into consideration the possible short generation times in primate ancestors, it is concluded that the ancestral population size in the primate lineage was no smaller than that of extant humans. Furthermore, under the assumption of 6 million years (myr) divergence between humans and chimpanzees, the divergence time of humans from gorillas, orangutans, Old World monkeys, and New World monkeys is estimated as 7.2, 18, 34, and 65 myr ago, respectively, which are generally older than traditional estimates. Beside the intron sequences, three other data sets of orthologous sequences are used between the human and the chimpanzee comparison. The ML application to these data sets including 58,156 random BAC end sequences (BES) shows that the nucleotide substitution rate is as low as 0.6–0.8 × 10^–9 per site per year and the extent of ancestral polymorphism is 0.33–0.51%. With such a low substitution rate and short generation time, the relatively high extent of polymorphism suggests a fairly large effective population size in the ancestral lineage common to humans and chimpanzees.[Reviewing Editor: Dr. Magnus Nordborg]     

ASPM and the Evolution of Cerebral Cortical Size in a Community of New World Monkeys

The ASPM (abnormal spindle-like microcephaly associated) gene has been proposed as a major determinant of cerebral cortical size among primates, including humans. Yet the specific functions of ASPM and its connection to human intelligence remain controversial. This debate is limited in part by a taxonomic focus on Old World monkeys and apes. Here we expand the comparative context of ASPM sequence analyses with a study of New World monkeys, a radiation of primates in which enlarged brain size has evolved in parallel in spider monkeys (genus Ateles) and capuchins (genus Cebus). The primate community of Costa Rica is perhaps a model system because it allows for independent pairwise comparisons of smaller- and larger-brained species within two taxonomic families. Accordingly, we analyzed the complete sequence of exon 18 of ASPM in Ateles geoffroyi, Alouatta palliata, Cebus capucinus, and Saimiri oerstedii. As the analysis of multiple species in a genus improves phylogenetic reconstruction, we also analyzed eleven published sequences from other New World monkeys. Our exon-wide, lineage-specific analysis of eleven genera and the ratio of rates of nonsynonymous to synonymous substitutions (d_N/d_S) on ASPM revealed no detectable evidence for positive selection in the lineages leading to Ateles or Cebus, as indicated by d_N/d_S ratios of <1.0 (0.6502 and 0.4268, respectively). Our results suggest that a multitude of interacting genes have driven the evolution of larger brains among primates, with different genes involved in this process in different encephalized lineages, or at least with evidence for positive selection not readily apparent for the same genes in all lineages. The primate community of Costa Rica may serve as a model system for future studies that aim to elucidate the molecular mechanisms underlying cognitive capacity and cortical size.     

Loss of olfactory receptor genes coincides with the acquisition of full trichromatic vision in primates

Olfactory receptor (OR) genes constitute the molecular basis for the sense of smell and are encoded by the largest gene family in mammalian genomes. Previous studies suggested that the proportion of pseudogenes in the OR gene family is significantly larger in humans than in other apes and significantly larger in apes than in the mouse. To investigate the process of degeneration of the olfactory repertoire in primates, we estimated the proportion of OR pseudogenes in 19 primate species by surveying randomly chosen subsets of 100 OR genes from each species. We find that apes, Old World monkeys and one New World monkey, the howler monkey, have a significantly higher proportion of OR pseudogenes than do other New World monkeys or the lemur (a prosimian). Strikingly, the howler monkey is also the only New World monkey to possess full trichromatic vision, along with Old World monkeys and apes. Our findings suggest that the deterioration of the olfactory repertoire occurred concomitant with the acquisition of full trichromatic color vision in primates.     

Genetic evidence of a strong functional constraint of neurotrypsin during primate evolution

Neurotrypsin is one of the extra-cellular serine proteases that are predominantly expressed in the brain and involved in neuronal development and function. Mutations in humans are associated with autosomal recessive non-syndromic mental retardation (MR). We studied the molecular evolution of neurotrypsin by sequencing the coding region of neurotrypsin in 11 representative non-human primate species covering great apes, lesser apes, Old World monkeys and New World monkeys. Our results demonstrated a strong functional constraint of neurotrypsin that was caused by strong purifying selection during primate evolution, an implication of an essential functional role of neurotrypsin in primate cognition. Further analysis indicated that the purifying selection was in fact acting on the SRCR domains of neurotrypsin, which mediate the binding activity of neurotrypsin to cell surface or extra-cellular proteins. In addition, by comparing primates with three other mammalian orders, we demonstrated that the absence of the first copy of the SRCR domain (exon 2 and 3) in mouse and rat was due to the deletion of this segment in the murine lineage.     

Electrophoretic polymorphism of proteins and the genetic divergence of primates

The hypothesis suggesting that genetic distances between primate taxa are smaller than characteristic genetic distances between non-primate taxa having the similar level of phylogenetic affinity, due to the specific features of primate protein evolution, has been probed. To this end, genetic distances between green and rhesus monkeys representing different genera of one subfamily, and between humans and chimpanzees representing related families, have been calculated and compared. It has been shown that the former are 2-2,5 times smaller than the latter. It is pointed out in this connection that genetic distances reflect adequately the hierarchy of the above taxa, and the existing interpretation of the "paradox of genetic similarity" of man and apes needs to be corrected. To calculate genetic distances, we used both literature data and the results of comparative analysis of 9 electrophoretic gene markers of green and rhesus monkeys represented in this work. Differences in genetic variability of these species were detected.     

Sequences from the 5′ flanking region of the ϵ‐globin gene support the relationship of Callicebus with the pitheciins

The purpose of this study was to determine nucleotide sequences from the 5′ flanking region of the ϵ‐globin gene of selected platyrrhine primates and to analyze the data for phylogenetic information and estimated times of divergence. We report new sequence data for two species of New World monkeys, Callicebus torquatus and Pithecia irrorata. We analyzed these data in conjunction with homologous sequences from other primate species. The data support the hypothesis that the titi monkeys (Callicebus) and seed predators (Tribe Pitheciini) form a clade (Subfamily Pitheciinae), and also provide limited support for that subfamily being allied with the atelines. We also present estimated dates of divergence for the Callicebus and pitheciin lineages. Am. J. Primatol. 48:69–75, 1999. © 1999 Wiley‐Liss, Inc.     

Mhc-DQB repertoire variation in hominoid and Old World primate species.

Comparison of 87 distinct Mhc-DQB sequences, obtained from 13 primate species, demonstrates that five out of eight trans-species Mhc-DQB allele lineages are at least 30 million years old and predate divergence of hominoid and Old World primate species. One lineage may be much older because its members are not only traced back in higher primates, but also are present in a New World primate species. Comparing Mhc-DQB repertoire variation in distinct species, allows one to pinpoint when certain polymorphisms were lost or gained in primate evolution. Heterogeneity observed among members of trans-species Mhc-DQB allele lineages can be explained in major part by point mutations, whereas intraexonic crossing-over is a potent mechanism in generating new allele lineages. The stability of Mhc-DQB polymorphisms is influenced by selective forces because distinct allele lineages appear to have accumulated nucleotide substitutions and amino acid replacements at different rates.     

Molecular evolution of the psi eta-globin gene locus: gibbon phylogeny and the hominoid slowdown

An 8.4-kb genomic region spanning both the psi eta-globin gene locus and flanking DNA was sequenced from the common gibbon (Hylobates lar). In addition, sequencing of the entire orthologous region from galago (Galago crassicaudatus) was completed. The gibbon and galago sequences, along with published orthologous sequences from 10 other species, were aligned. These noncoding nucleotide sequences represented four human alleles, four apes (chimpanzee, gorilla, organgutan, and gibbon), an Old World monkey (rhesus monkey), two New World monkeys (spider and owl monkeys), tarsier, two strepsirhines (galago and lemur), and goat. Divergence and maximum parsimony analyses of the psi eta genomic region first groups humans and chimpanzees and then, at progressively more ancient branch points, successively joins gorillas, orangutans, gibbons, Old World monkeys, New World monkeys, tarsiers, and strepsirhines (the lemuriform-lorisiform branch of primates). This cladistic pattern supports the taxonomic grouping of all extant hominoids into family Hominidae, the division of Hominidae into subfamilies Hylobatinae (gibbons) and Homininae, the division of Homininae into tribes Pongini (orangutans) and Hominini, and the division of Hominini into subtribes Gorillina (gorillas) and Hominina (chimpanzees and humans). The additional gibbon and galago sequence data provide further support for the occurrence of a graded evolutionary-rate slowdown in the descent of simian primates, with the slowing rate being more pronounced in the great-ape and human lineages than in the gibbon or monkey lineages. A comparison of global versus local molecular clocks reveals that local clock predictions, when focused on a specific number of species within a narrow time frame, provide a more accurate estimate of divergence dates than do those of global clocks.     

β-Defensin 1 gene variability among non-human primates

Defensins are a recently described family of peptides that play an important role in innate immunity. Recent studies have shown that defensins exhibit a broad spectrum of antimicrobial activities against bacteria and fungi. Three families have been identified so far in mammals, alpha-defensins, beta-defensins and theta-defensins, presumably derived from a common ancestral defensin. A long-term study on the evolution of these multigene families among primates has been undertaken to investigate: (1) the degree of interspecific differentiation; (2) the genetic mechanisms responsible for the variability of these molecules; and (3) the possible role of different environmental factors in their evolution. Nucleotide sequences have been obtained from great and lesser apes, several African and Asian catarrhine monkeys and one New World monkey. A comparison of rates of synonymous and nonsynonymous (amino-acid changing) nucleotide substitution indicates that the primate beta-defensin 1 gene evolved under a pattern of random nucleotide substitution as predicted by the neutral theory of molecular evolution. These results are not consistent with the hypothesis that the primate beta-defensin 1 gene has diversified in response to changes in the microbial species to which a given host is exposed. Analyses of interpecific variability have yielded some insights about the pattern of molecular evolution of the gene among primates. Humans and great apes present high levels of sequence similarity, differing in only one amino acid residue in the mature peptide. Compared with these taxa, hylobatids and cercopithecids exhibit 3-4 amino acid substitutions, some of which increase the net charge of the active molecule.     

Direct estimate of the spontaneous germ line mutation rate in African green monkeys

Here, I provide the first direct estimate of the spontaneous mutation rate in an Old World monkey, using a seven individual, three‐generation pedigree of African green monkeys. Eight de novo mutations were identified within ～1.5 Gbp of accessible genome, corresponding to an estimated point mutation rate of 0.94 × 10^?8 per site per generation, suggesting an effective population size of ～12000 for the species. This estimation represents a significant improvement in our knowledge of the population genetics of the African green monkey, one of the most important nonhuman primate models in biomedical research. Furthermore, by comparing mutation rates in Old World monkeys with the only other direct estimates in primates to date–humans and chimpanzees–it is possible to uniquely address how mutation rates have evolved over longer time scales. While the estimated spontaneous mutation rate for African green monkeys is slightly lower than the rate of 1.2 × 10^?8 per base pair per generation reported in chimpanzees, it is similar to the lower range of rates of 0.96 × 10^?8–1.28 × 10^?8 per base pair per generation recently estimated from whole genome pedigrees in humans. This result suggests a long‐term constraint on mutation rate that is quite different from similar evidence pertaining to recombination rate evolution in primates.     

Presence and abundance of CENP-B box sequences in great ape subsets of primate-specific α-satellite DNA

CENP-B, a highly conserved centromere-associated protein, binds to -satellite DNA, the centromeric satellite of primate chromosomes, at a 17-bp sequence, the CENP-B box. By fluorescence in situ hybridization (FISH) with an oligomer specific for the CENP-B box sequence, we have demonstrated the abundance of CENP-B boxes on all chromosomes (except the Y) of humans, chimpanzee, pygmy chimpanzee, gorilla, and orangutan. This sequence motif was not detected in the genomes of other primates, including gibbons, Old and New World monkeys, and prosimians. Our results indicate that the CENP-B box containing subtype of -satellite DNA may have emerged recently in the evolution of the large-bodied hominoids, after divergence of the phylogenetic lines leading to gibbons and apes; the box is thus on the order of 15–25 million years of age. The rapid process of dispersal and fixation of the CENP-B box sequence throughout the human and great ape genomes is thought to be a consequence of concerted evolution of -satellite subsets on both homologous and nonhomologous chromosomes.Correspondence to: T. Haaf     

Molecular evidence on Primate phylogeny from DNA sequences

Evidence from DNA sequences on the phylogenetic systematics of primates is congruent with the evidence from morphology in grouping Cercopithecoidea (Old World monkeys) and Hominoidea (apes and humans) into Catarrhini, Catarrhini and Platyrrhini (ceboids or New World monkeys) into Anthropoidea, Lemuriformes and Lorisiformes into Strepsirhini, and Anthropoidea, Tarsioidea, and Strepsirhini into Primates. With regard to the problematic relationships of Tarsioidea, DNA sequences group it with Anthropoidea into Haplorhini. In addition, the DNA evidence favors retaining Cheirogaleidae within Lemuriformes in contrast to some morphological studies that favor placing Cheirogaleids in Lorisiformes. While parsimony analysis of the present DNA sequence data provides only modest support for Haplorhini as a monophyletic taxon, it provides very strong support for Hominoidea, Catarrhini, Anthropoidea, and Strepsirhini as monophyletic taxa. The parsimony DNA evidence also rejects the hypothesis that megabats are the sister group of either Primates or Dermoptera (flying lemur) or a Primate-Dermoptera clade and instead strongly supports the monophyly of Chiroptera, with megabats grouping with microbats at considerable distance from Primates. In contrast to the confused morphological picture of sister group relationships within Hominoidea, orthologous noncoding DNA sequences (spanning alignments involving as many as 20,000 base positions) now provide by the parsimony criterion highly significant evidence for the sister group relationships defined by a cladistic classification that groups the lineages to all extant hominoids into family Hominidae, divides this ape family into subfamilies Hylobatinae (gibbons) and Homininae, divides Homininae into tribes Pongini (orangutans) and Hominini, and divides Hominini into subtribes Gorillina (gorillas) and Hominina (humans and chimpanzees). A likelihood analysis of the largest body of these noncoding orthologues and counts of putative synapomorphies using the full range of sequence data from mitochondrial and nuclear genomes also find that humans and chimpanzees share the longest common ancestry. © 1994 Wiley-Liss, Inc.     

SINE insertions in cladistic analyses and the phylogenetic affiliations of Tarsius bancanus to other primates

Transpositions of Alu sequences, representing the most abundant primate short interspersed elements (SINE), were evaluated as molecular cladistic markers to analyze the phylogenetic affiliations among the primate infraorders. Altogether 118 human loci, containing intronic Alu elements, were PCR analyzed for the presence of Alu sequences at orthologous sites in each of two strepsirhine, New World and Old World monkey species, Tarsius bancanus, and a nonprimate outgroup. Fourteen size-polymorphic amplification patterns exhibited longer fragments for the anthropoids (New World and Old World monkeys) and T. bancanus whereas shorter fragments were detected for the strepsirhines and the outgroup. From these, subsequent sequence analyses revealed three Alu transpositions, which can be regarded as shared derived molecular characters linking tarsiers and anthropoid primates. Concerning the other loci, scenarios are represented in which different SINE transpositions occurred independently in the same intron on the lineages leading both to the common ancestor of anthropoids and to T. bancanus, albeit at different nucleotide positions. Our results demonstrate the efficiency and possible pitfalls of SINE transpositions used as molecular cladistic markers in tracing back a divergence point in primate evolution over 40 million years old. The three Alu insertions characterized underpin the monophyly of haplorhine primates (Anthropoidea and Tarsioidea) from a novel perspective.