*Department of Biosystems Science, Graduate University for Advanced Studies (Sokendai), Hayama, Kanagawa, Japan;
Primate Research Institute, Kyoto University, Inuyama, Aichi, Japan
![]() |
Abstract |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
![]() |
Introduction |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
It has been proposed that because uric acid is a powerful scavenger of free radicals, it plays an important role in protecting hominoids from oxidative damage (Ames et al. 1981
; Whiteman and Halliwell 1996
). Recently, Scott and Hooper (2001)
have further argued that the accumulation of uric acid in hominoids results from its capability to inhibit the peroxynitrite-dependent extravasation of inflammatory cells and thereby prevent oxidative damage to the increasingly complex brains. Another interesting postulate, which Haldane (1955)
questioned, though, was made by Orowan (1955)
on the basis of the structural similarity of uric acid to cerebral stimulants, caffeine and theobromine. According to this idea, loss of Uox activity might result in a quantum jump in intellectual capability and thus trigger emergence of man.
Wu et al. (1989, 1992)
first isolated the human genomic DNA containing the Uox gene and sequenced the whole coding region. In addition, they examined the chimpanzee, orangutan, and gibbon orthologs regarding the presence or absence of three detrimental mutations that were found in the human (two nonsense mutations at codons 33 and 187 and one mutant splice acceptor signal of exon 3). They found that the nonsense mutation at codon 33 was shared by the chimpanzee and the orangutan but that at codon 187 existed only in the chimpanzee. Moreover, because none of these three detrimental mutations was present in the gibbon, they partially sequenced the gibbon exon 2 and found a 13-bp deletion between codons 72 and 76. On the basis of these findings, they came to the conclusion that two independent inactivations of the Uox gene occurred in hominoids: the nonsense mutation at codon 33 in the human, the chimpanzee, and the orangutan (see also Yeldandi et al. 1991
) and the 13-bp deletion in the gibbon. However, these studies focused on exons 2 and 5 only in rather limited primate species.
We therefore decided to fully explore the evolutionary change of the Uox gene in five hominoids, three Old World monkeys (Catarrhini), and one New World monkey (Platyrrhini). We determined the DNA sequences of the entire coding region of 915 bp for all these nine species. We also sequenced the promoter region of about 1,400 bp to examine if the Uox gene was silenced progressively or suddenly and sequenced intron 4 of about 1,400 bp to measure relative selective constraint against the promoter and coding regions. Furthermore, to see if the 13-bp deletion is common among gibbons, we sequenced exons 25 for three subgenera of the family Hylobatidae. In this paper, we report comparative analyses of these sequences and discuss the evolutionary implications of loss of Uox activity in hominoids.
![]() |
Materials and Methods |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
PCR Amplification, Cloning, and Sequencing
For the human, chimpanzee, gorilla, orangutan, white-handed gibbon, baboon, rhesus monkey, crab-eating monkey, and owl monkey Uox genes, the entire eight exons of 915 bp, the promoter region of ca. 1,400 bp upstream of the initiation codon, and part of intron 4 of ca. 1,400 bp were amplified by polymerase chain reaction (PCR). In order to examine the inactivation process of the Uox gene in the family Hylobatidae, similar PCR amplification was also carried out for exons 25 for another H. lar, as well as for H. agilis, H. muelleri, H. concolor, and H. syndactylus. On the basis of available human Uox genomic sequences (GenBank accession number S94095 and AL136113), PCR primers for each region were designed. Each PCR reaction was done in a 25-µl volume. The reaction mixture contained 50100 ng of genomic DNA, 10 mM Tris-HCl (pH 8.3), 50 mM KCl, 1.5 mM MgCl2, 0.2 mM of dNTPs, 1.5 U of TaKaRa EX TaqTM polymerase (TaKaRa), and 0.4 µM of the upper and lower primer. PCR primer sequences and conditions were adapted to each region and are available upon request. All PCR products were purified through QIAquick PCR Purification Kit (QIAGEN). For exons 3, 4, and 68 in the nine primates and exons 25 in additional gibbons, PCR products were directly sequenced. Other PCR products of the promoter region, exons 1, 2, and 5, and intron 4 were cloned into pCRII-TOPO through TOPO TA Cloning Kits (Invitrogen). Plasmid DNAs were purified with QIAprep Spin Miniprep (QIAGEN) and used as templates in sequencing reaction. Sequencing reaction was performed with ABI prism Dye-Deoxy Reaction Kit (PE Applied Biosystems) and analyzed on ABI PRISM 377 DNA sequencer (PE Applied Biosystems). To avoid sequencing errors, PCR products or plasmid DNAs were sequenced two to six times for each product or plasmid in both directions. Sequences were also confirmed by independent PCRs. These sequences were deposited in the DNA Data Bank of Japan (DDBJ), and their accession numbers are given in Appendix I.
Data Analysis
DNA sequence alignment with insertion of gaps was performed with CLUSTAL W (Thompson, Higgins, and Gibson 1994
), and the aligned sequences were manually checked by eye (Appendix II). In the phylogenetic analysis, any site containing insertion or deletion (indel) was excluded. We used two tree-making methods: neighbor-joining (NJ) by Saitou and Nei (1987)
and maximum parsimony (MP) by Fitch (1971)
and Hartigan (1973)
in PHYLIP version 3.57c (Felsenstein 1995
). For the coding region, the synonymous and nonsynonymous sites and the numbers of nucleotide differences at these respective sites were counted by the method of Nei and Gojobori (1986)
and by the modified version with transition bias (R = 1.0) in Nei and Kumar (2000
, pp. 5759). When necessary, multiple-hit substitutions were corrected based on either Kimura's two-parameter model (Kimura 1980, 1983
, pp. 9097) or JukesCantor model (Jukes and Cantor 1969
).
![]() |
Results |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
|
In the five hominoid -Uox genes, there are several independent nonsense mutations in five CGA, one TGG, or one CAG codon (Appendix II). Furthermore, there exists one mutation in the splice acceptor signal of exon 3 shared by the human and the African apes (data not shown), one human-specific 26-bp deletion in the enhancer region, one gorilla-specific mutation in the initiation codon, and one gorilla-specific tandem duplication in exon 2. Of these nonsense mutations, three occur in species-specific manners, and only one at codon 33 in exon 2 is shared by the human and the great apes. As previously inferred by Wu et al. (1992)
, it is therefore reasonable to conclude that the
-Uox gene in the human and the great apes was caused by this shared nonsense mutation. Another nonsense mutation at codon 187 is shared by the human, the chimpanzee and the gorilla, so that it is most likely to have occurred in their ancestral species. The nonsense mutation (TGA) at codon 107 is, however, more complicated than others. It occurs in the gorilla, the orangutan, and the gibbon, and therefore requires multiple origins of this nonsense mutation. Overall, it is remarkable that, except one CGA codon in exon 6, all the other four CGA codons are converted to the TGA termination codon in all or some of the hominoids. Arginine (Arg) is a basic amino acid, and its usage varies greatly from gene to gene. However, in most organisms, Arg is generally coded by non-CGA codons (Wada et al. 1992
; Osawa 1995
, pp. 4557). The fact that there are as many as five CGA among 12 Arg codons in the Old World monkey Uox is suggestive of the high likelihood of eventual dysfunctioning of this gene.
To obtain g in -Uox gene lineages, we use the MP estimates for all branches that occur in the clade of the human and the great apes (table 2
). The gibbon Uox is also nonfunctional, but we exclude it from the present calculation because its dysfunctioning occurred independently in that lineage and its timing is unknown (Wu et al. 1992
). In total, there are 68.5 nucleotide substitutions in intron 4, 58.7 in the promoter region, and 25.5 at the nonsynonymous sites. We therefore have gp = (58.7/1340)/(68.5/1336) = 0.85 for the former and gn = (25.5/659)/(68.5/1336) = 0.75 for the latter. At face value, both gp and gn of
-Uox are smaller than 1. However, it is difficult to imagine any selective constraint against pseudogenes; in fact, the result of the G-test shows that the observed gp = 0.86 and gn = 0.75 values are not significantly different from 1 (df = 1, P > 0.1 in both cases). We thus conclude the absence of selective constraint against the promoter region (but see Discussion) as well as the nonsynonymous sites in the
-Uox gene lineage and subsequently assume that g = 1 in both regions.
|
Dating the Inactivation Event
A dramatic change in the degree of functional constraint at the nonsynonymous sites can be used to estimate when the Uox gene became nonfunctional. We separately treat two independent events of gene dysfunctioning in the hominoid lineage: one in the stem lineage of the human and the great apes and the other in the stem lineage of gibbons. We designate by x the time period during which the gene was functional since it diverged from the Catarrhini lineage and by y the time period of inactivation in the lineage leading to the human and the great apes or to the gibbon (fig. 2
). As mentioned previously, the proportion of selective neutrality during these two time periods is given by fn and gn, respectively. If we denote the per-site substitution rate in intron 4 by k per unit time, we have
|
|
|
In the case of the gibbon lineage, the MP estimates of a and b become 0.033 and 0.014, respectively. The height of node B relative to node A becomes 24.3/44.6 = 0.54 in intron 4. Again using the same estimates of fn and gn as before, we have r = 0.28 estimated from (2). This r value is considerably smaller than the relative height of node B and consistent with the finding that the gibbon Uox gene was inactivated independently of the human and great ape lineage (Wu et al. 1992
). Although most species divergence times in primate phylogeny are still controversial (Martin 1993
; Kumar and Hedges 1998
; Takahata 2001
), it is interesting to convert these ratios of r into the geological times. If we assume that Catarrhini and hominoids diverged 35 MYA, then r = 0.44 and 0.28 correspond to 15.4 and 9.8 MYA, respectively. Importantly, this relatively recent inactivation in the gibbon lineage suggests that some species in the family Hylobatidae may still possess the functional Uox gene. Although Friedman et al. (1985)
were unable to detect Uox activity in the adults of H. lar and H. concolor, there is at present no information about subgenera Bunopithecus and Symphalangus. In order to examine this possibility and which mutation was responsible for the dysfunctioning of gibbon
-Uox, we have examined exon 25 DNA sequences for several gibbon species.
Inactivation in the Gibbon Lineage
There are seven seemingly detrimental mutations in the gibbon -Uox gene: two in exon 2, three in exon 3, one in the splice donor site at intron 3, and one in exon 5. One such is the 13-bp deletion in exon 2, which is previously thought to be responsible for the dysfunctioning of the gibbon Uox gene (Wu et al. 1992
). In the present study, we have newly examined H. agilis, H. muelleri, H. concolor, and H. syndactylus in addition to two individuals of H. lar. The 13-bp deletion is indeed shown to be present in subgenus Hylobates (2n = 44) including H. lar, H. agilis, and H. muelleri. However, neither subgenus Nomascus (2n = 52) including H. concolor nor subgenus Symphalangus (2n = 50) including H. syndactylus possesses this deletion, and these subgenera exhibit the same stretch of AGAACACAGTTCA as in the human and the great apes. A parsimonious explanation is that the deletion took place in the common ancestral lineage of subgenus Hylobates. According to the recent study on the major Hylobatid divisions by Roos and Geissmann (2001)
, H. nomascus branched off first, followed by Symphalangus, and then by Bunopithecus and Hylobates. The Uox sequence differences between Symphalangus and Hylobates are smaller than those between the human and the orangutan, suggesting the relatively recent event of the 13-bp deletion.
One nonsense mutation (CGATGA) is found in each of exon 2 and exon 3 (fig. 3
). It turns out that the exon 2 mutation is shared by all gibbon species examined and is not found in any of the remaining primate species. It is therefore most likely that this mutation took place before the subgenus differentiation and inactivated the gibbon Uox gene altogether. If this is the case and if the inactivation resulting from the nonsense mutation occurred about 10 MYA as mentioned previously, the subgenus differentiation in extant gibbons must have taken place during the past 10 Myr (Roos and Geissmann 2001
and references therein). In contrast, the exon 3 mutation is not shared by H. syndactylus but by the gorilla and the orangutan. The origin of this mutation is therefore multiple and relatively recent in the gibbon lineage. The splice donor site mutation at intron 3 is shared by gibbons, except H. syndactylus (data not shown). This and other substitutions in the gibbon Uox gene support that H. syndactylus is the most distantly related species in the family (fig. 3
). This conclusion is different from that of Roos and Geissman (2001)
, who studied the mitochondrial control region and Phe-tRNA. In addition to nonsense and splice donor site mutations, there are several indels. One such is a two-base deletion in exon 3 which occurs specifically to subgenus Hylobates. There are also one-base deletion in exon 3 and one-base insertion in exon 5, both of which are specific to all the gibbon lineages. The frameshift caused by the one-base deletion produces a termination codon in exon 4 and that caused by the one-base insertion produces a termination codon within exon 5. Because the active center of the enzyme is coded by codon 127, 129, and 131 in exon 4 (Wu et al. 1989
; Chu et al. 1996
), the one-base deletion may be more detrimental than the one-base insertion. However, this information alone seems insufficient to exclude the one-base insertion from the cause of the gene inactivation.
|
![]() |
Discussion |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
It has been recently reported that the Uox gene and the DLAD (DNase II-Like Acid DNase) gene lie in a head-to-head orientation (Shiokawa and Tanuma 2001)
and that exon 1 followed by large intron 1 of the DLAD gene is located from -776 to -900 bp in the promoter region of the Uox gene (Appendix II). Because of this, the promoter region of the DLAD gene, though not yet well characterized, should overlap with that of the Uox gene. Because the DLAD gene is presumably functional in the hominoids (Shiokawa and Tanuma 2001
), this overlap can explain why the promoter region of the
-Uox gene in the hominoids is not completely free from selective constraint (table 2
). The fact that the constraint in the region proximal to the initiation codon of the
-Uox gene (-1 to -400 bp) is more apparent than in the distal region (-401 to -700 bp) also suggests that this proximal region still plays important roles in DLAD transcription.
Loss of Uox activity in hominoids results in >10-fold higher concentration of uric acid in blood of humans and most primates than in other mammals (Keilin 1959
). Although uric acid as a powerful antioxidant may provide a basis for the increase of life span and decrease cancer rates (Ames et al. 1981
), the high concentration predisposes to crystalline deposition (Slot 1994
). One way to reduce the risk is to decrease the xanthine oxidoreductase (Xor) gene activity (Xu, LaVallee, and Hoidal 2000
). It is known that the Xor enzymatic activity in humans is 100 times lower than that in bovine, rats, and mice (Abadeh et al. 1992
). Xu, LaVallee, and Hoidal (2000)
demonstrated that transcription and core promoter activity of the human Xor are repressed. It appears that the purine metabolic system in humans has found a way to avoid the overproduction of uric acid by downregulating the Xor gene expression, the product converting hypoxanthine to xanthine and further xanthine to uric acid. However, it is not known how and when this coevolution began. If the Uox gene expression has been repressed since the early evolution of primates, the repression of Xor gene expression might also be favored by natural selection from that time on. On the other hand, if the downregulation of Xor gene expression is hominoid-specific, the coevolution must be relatively recent. In this case, however, there is no reason to think that both great and lesser apes could downregulate the Xor gene expression in the same way. Whichever is the case, the purine metabolic system should provide an excellent example for studying molecular coevolution, and examination of the Xor regulatory elements in nonhuman primates will be of particular interest in this regard.
|
![]() |
Appendix II |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Dashes () indicate identity with top sequence, and asterisks (*) indicate postulated deletions. The promoter region, coding region, and intron 4 are presented in this order separately. In the promoter region, the rat ortholog (D50689) is also aligned from the initiation codon to -320 bp; the enhancer region is underlined, the 5' untranslated region is doubly underlined, and the cis-acting element, palindromic sequence, and CAAT-box are boxed. In the coding region, the mutation in the initiation codon in the gorilla is shaded, and the termination codons produced by nonsense mutations are boxed. Three TGA nonsense mutations at codon 107 appear to occur independently in the gorilla, the orangutan, and the gibbon. However, in the oraguntan, codon 107 is polymorphic with respect to TGA and CGA, whereas in the H. syndactylus, codon 107 is occupied by CGA.
|
|
|
![]() |
Acknowledgements |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
![]() |
Footnotes |
---|
Abbreviations: Uox, urate oxidase; Xor, xanthine oxidoreductase; Hosa, Homo sapiens; Patr, Pan troglodytes; Gogo, Gorilla gorilla; Popy, Pongo pygmaeus; Hyla, Hylobates lar; Hyag, H. agilis; Hymu, H. muelleri; Hyco, H. concolor; Hysy, H. syndactylus; Paha, Papio hamadryas; Mamu, Macaca mulatta; Mafa, M. fascicularis; Aotr, Aotus trivirgatus.
Keywords: uric acid
purine metabolism
primates
codon bias
Address for correspondence and reprints: Naoyuki Takahata, Department of Biosystems Science, Graduate University for Advanced Studies, Hayama, Kanagawa 240-0193, Japan. takahata{at}soken.ac.jp
.
![]() |
References |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Abadeh S., J. Killachy, M. Benboubetra, R. Harrison, 1992 Purification and partial characterization of xanthine oxidase from human milk Biochim. Biophys. Acta 1117:25-32[ISI][Medline]
Ames B. N., R. Cathcart, E. Schwiers, E. Hochstein, 1981 Uric acid provides an antioxidant defense in humans against oxidant- and radical-caused aging and cancer: a hypothesis Proc. Natl. Acad. Sci. USA 78:6858-6862[Abstract]
Chen F.-C., W.-H. Li, 2001 Genomic divergences between humans and other hominoids and the effective population size of the common ancestor of humans and chimpanzees Am. J. Hum. Genet 68:444-456[ISI][Medline]
Cheng X. G., M. Nomura, K. Takane, H. Kouchi, S. Tajima, 2000 Expression of two uricase (Nodulin-35) genes in a non-ureide type legume, Medicago sativa Plant Cell Physiol 41:104-109[ISI][Medline]
Christen P., W. C. Peacock, A. E. Christen, W. E. C. Wacker, 1970a. Urate oxidase in primate phylogenesis Eur. J. Biochem 12:3-5[ISI][Medline]
. 1970b. Urate oxidase in primates Folia Primatol 13:35-39[ISI][Medline]
Chu R., Y. Lin, N. Usuda, M. S. Rao, J. K. Reddy, A. V. Yeldandi, 1996 Mutational analysis of the putative copper-binding site of rat urate oxidase Ann. N.Y. Acad. Sci 27:781-786
Felsenstein J., 1995 PHYLIP (phylogeny inference package) Version 3.57c. University of Washington, Seattle
Fitch W. M., 1971 Toward defining the course of evolution: minimum change for a specific tree topology Syst. Zool 20:406-416[ISI]
Friedman T. B., G. E. Polanco, J. C. Appold, J. E. Mayle, 1985 On the loss of uricolytic activity during primate evolutionI. Silencing of urate oxidase in a hominoid ancestor Comp. Biochem. Physiol 81B:653-659
Fujiwara S., K. Nakashima, T. Noguchi, 1987 Insoluble uricase in liver peroxisomes of old world monkeys Comp. Biochem. Physiol 88B:467-469[ISI]
Haldane J. B. S., 1955 Origin of man Nature 176:169-170[ISI][Medline]
Hartigan J. A., 1973 Minimum evolution fits to a given tree Biometrics 29:53-65[ISI]
Izuhara M., M. Ito, Y. Takagi, 1995 Transcription of the rat liver uricase-encoding gene is regulated via a cis-acting element responsive to cAMP Gene 167:267-272[ISI][Medline]
Jukes T. H., C. R. Cantor, 1969 Evolution of protein molecules Pp. 21132 in H. N. Munro, ed. Mammalian protein metabolism. Academic Press, New York
Karlin S., C. Burge, 1995 Dinucleotide relative abundance extremes: a genome signature Trends Genet 11:283-290[ISI][Medline]
Keilin J., 1959 The biological significance of uric acid and guanine excretion Biol. Rev 34:265-296[ISI]
Kimura M., 1980 A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences J. Mol. Evol 16:111-120[ISI][Medline]
. 1983 The neutral theory of molecular evolution Cambridge University Press, Cambridge
Kumar S., S. B. Hedges, 1998 A molecular timescale for vertebrate evolution Nature 392:917-919[ISI][Medline]
Lehninger A. L., 1995 Biochemistry 2nd edition. Kalyani Publications, Ludhiana, New Delhi
Logan D. C., D. E. Wilson, C. M. Flowers, P. J. Sparks, F. H. Tyler, 1976 Uric acid catabolism in the woolly monkey Metabolism 25:517-522[ISI][Medline]
Lootens S., J. Burnett, T. B. Friedman, 1993 An intraspecific gene duplication polymorphism of the urate oxidase gene of Drosophila virilis: a genetic and molecular analysis Mol. Biol. Evol 10:635-646[Abstract]
Martin R. D., 1993 Primate origins, plugging the gaps Nature 356:121-125[ISI]
Nei M., T. Gojobori, 1986 Simple methods for estimating the numbers of synonymous and nonsynonymous nucleotide substitutions Mol. Biol. Evol 3:418-426[Abstract]
Nei M., S. Kumar, 2000 Molecular evolution and phylogenetics Oxford University Press, New York
Ohno S., 1988 Universal rule for coding sequence construction: TA/CG deficiency-TG/CT excess Proc. Natl. Acad. Sci. USA 85:9630-9634[Abstract]
Orowan E., 1955 The origin of man Nature 175:683-684[ISI][Medline]
Osawa S., 1995 Evolution of the genetic code Oxford University Press, Tokyo
Roos C., T. Geissmann, 2001 Molecular phylogeny of the major hylobatid divisions Mol. Phylogenet. Evol 19:486-494[ISI][Medline]
Saitou N., M. Nei, 1987 The neighbor-joining method: a new method for reconstructing phylogenetic trees Mol. Biol. Evol 4:406-425[Abstract]
Satta Y., J. Klein, N. Takahata, 2000 DNA archives and our nearest relative: the trichotomy problem revisited Mol. Phylogenet. Evol 14:259-275[ISI][Medline]
Scott G. S., D. C. Hooper, 2001 The role of uric acid in protection against peroxynitrite-mediated pathology Med. Hypotheses 56:95-100[ISI][Medline]
Shioiri C., N. Takahata, 2001 Skew of mononucleotide frequencies, relative abundance of dinucleotides, and DNA strand asymmetry J. Mol. Evol 53:364-376[ISI][Medline]
Shiokawa D., S. Tanuma, 2001 Isolation and characterization of the DLAD/Dlad genes, which lie head-to-head with the genes for urate oxidase Biochem. Biophys. Res. Commun 288:1119-1128[ISI][Medline]
Simkin P. A., 1971 Uric acid metabolism in Cebus monkeys Am. J. Physiol 221:1105-1109
Slot O., 1994 Hyperuricemia Ugeskr Laeger 156:2396-2401[Medline]
Takahata N., 2001 Molecular phylogeny and demographic history of humans Pp. 299305 in P. V. Tobias, M. A. Raath, J. Moggi-Cecchi, and G. A. Doyle, eds. Humanity from African naissance to coming millenia. Firenze University Press, Firenze/Witwatersrand University Press, Johannesburg
Takane K., S. Tajima, H. Kouchi, 1997 Two distinct uricase II (nodulin 35) genes are differentially expressed in soybean plants Mol. Plant Microbe Interact 10:735-741[ISI][Medline]
Thompson J., D. G. Higgins, T. Gibson, 1994 Clustal W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, positions-specific gap penalties and weight matrix choice Nucleic Acids Res 22:4673-4680[Abstract]
Whiteman M., B. Halliwell, 1996 Protection against peroxynitrite-dependent tyrosine nitration and 1-antiproteinase inactivation by ascorbic acid A comparison with other biological antioxidants. Free Radic. Res 25:275-283
Wada K., Y. Wada, F. Ishibashi, T. Gojobori, T. Ikemura, 1992 Codon usage tabulated from the GenBank genetic sequence data Nucleic Acids Res 20:2111-2118[ISI][Medline]
Wu X., C. C. Lee, D. M. Muzny, C. T. Caskey, 1989 Urate oxidase: primary structure and evolutionary implications Proc. Natl. Acad. Sci. USA 86:9412-9416[Abstract]
Wu X., D. M. Muzny, C. C. lee, C. T. Caskey, 1992 Two independent mutational events in the loss of urate oxidase during hominoid evolution J. Mol. Evol 34:78-84[ISI][Medline]
Wu X., M. Wakamiya, S. Vaishnav, R. Geske, C. Montgomery Jr., P. Jones, A. Bradley, C. T. Caskey, 1994 Hyperuricemia and urate nephropathy in urate oxidase-deficient mice Proc. Natl. Acad. Sci. USA 91:742-746[Abstract]
Xu P., P. Lavallee, J. R. Hoidal, 2000 Repressed expression of the human xanthine oxidoreductase gene J. Biol. Chem 275:5918-5926
Yeldandi A. V., R. Chu, J. Pan, Y. Zhu, N. Usuda, 1996 Peroxisomal purine metabolism Ann. N.Y. Acad. Sci 804:165-175[ISI][Medline]
Yeldandi A. V., V. Yeldandi, S. Kumar, C. V. N. Murthy, X. Wang, K. Alvares, M. S. Rao, J. K. Reddy, 1991 Molecular evolution of the urate oxidase-encoding gene in hominoid primates: nonsense mutations Gene 109:281-284[ISI][Medline]