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Origins, distribution and expression of the Duarte-2 (D2) allele of galactose-1-phosphate uridylyltransferase.

Carney AE, Sanders RD, Garza KR, McGaha LA, Bean LJ, Coffee BW, Thomas JW, Cutler DJ, Kurtkaya NL, Fridovich-Keil JL - Hum. Mol. Genet. (2009)

Bottom Line: Patients with Duarte galactosemia demonstrate reduced GALT activity and carry one profoundly impaired GALT allele (G) along with a second, partially impaired GALT allele (Duarte-2, D2).The p.N314D substitution, however, is not; it is found together with a silent polymorphism, p.L218(TTA), on functionally normal Duarte-1 alleles (D1, also called Los Angeles or LA alleles).Here we present allele-specific qRT-PCR evidence that D2 alleles express less mRNA in vivo than their wild-type counterparts; the difference is small but statistically significant.

View Article: PubMed Central - PubMed

Affiliation: Emory University, Atlanta, USA

ABSTRACT
Duarte galactosemia is a mild to asymptomatic condition that results from partial impairment of galactose-1-phosphate uridylyltransferase (GALT). Patients with Duarte galactosemia demonstrate reduced GALT activity and carry one profoundly impaired GALT allele (G) along with a second, partially impaired GALT allele (Duarte-2, D2). Molecular studies reveal at least five sequence changes on D2 alleles: a p.N314D missense substitution, three intronic base changes and a 4 bp deletion in the 5' proximal sequence. The four non-coding sequence changes are unique to D2. The p.N314D substitution, however, is not; it is found together with a silent polymorphism, p.L218(TTA), on functionally normal Duarte-1 alleles (D1, also called Los Angeles or LA alleles). The HapMap database reveals that p.N314D is a common human variant, and cross-species comparisons implicate D314 as the ancestral allele. The p.N314D substitution is also functionally neutral in mammalian cell and yeast expression studies. In contrast, the 4 bp 5' deletion characteristic of D2 alleles appears to be functionally impaired in reporter gene transfection studies. Here we present allele-specific qRT-PCR evidence that D2 alleles express less mRNA in vivo than their wild-type counterparts; the difference is small but statistically significant. Furthermore, we characterize the prevalence of the 4 bp deletion in GG, NN and DG populations; the deletion appears exclusive to D2 alleles. Combined, these data strongly implicate the 4 bp 5' deletion as a causal mutation in Duarte galactosemia and suggest that direct tests for this deletion, as proposed here, could enhance or supplant current tests, which define D2 alleles on the basis of the presence and absence of linked coding sequence polymorphisms.

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Proposed origins and relationship of the p.N314, D1 and D2 alleles of human GALT. The predicted ancestral human GALT allele carries the (GTCA)3 + p.L218(CTA) + p.D314 sequences found in other hominid species. The p.D314N substitution occurs early in human evolution, and the 4 bp 5′ deletion and p.L218(TTA) silent substitution occur later, on distinct branches of the tree. The three intronic base changes reported to exist in cis with D2 alleles [c.378-27G>C or IVS4-27G > C, c.508-24G > A or IVS5-24G > A and c.507 + 62G > A or IVS5-62G > A (20–22), not illustrated here] presumably occurred subsequent to, or concurrently with, the 4 bp 5′ deletion on the D2 branch of the tree.
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DDP080F1: Proposed origins and relationship of the p.N314, D1 and D2 alleles of human GALT. The predicted ancestral human GALT allele carries the (GTCA)3 + p.L218(CTA) + p.D314 sequences found in other hominid species. The p.D314N substitution occurs early in human evolution, and the 4 bp 5′ deletion and p.L218(TTA) silent substitution occur later, on distinct branches of the tree. The three intronic base changes reported to exist in cis with D2 alleles [c.378-27G>C or IVS4-27G > C, c.508-24G > A or IVS5-24G > A and c.507 + 62G > A or IVS5-62G > A (20–22), not illustrated here] presumably occurred subsequent to, or concurrently with, the 4 bp 5′ deletion on the D2 branch of the tree.

Mentions: As a first step toward understanding the origin and relationship of the p.N314D (rs2070074 at position 34,639,442 of chromosome 9 in NCBI Build 36.1) and 5′ variations in human GALT, we performed cross-species comparisons of the appropriate coding and non-coding sequences from representative human, non-human hominid (e.g. chimpanzee), non-hominid primate (e.g. macaque) and non-primate placental mammalian (e.g. mouse) species. All species examined, other than human, encoded D rather than N at position 314 (Table 1), strongly implicating D314 as the ancestral GALT allele; the variant that is predominant among modern humans is therefore most appropriately termed p.D314 N (Fig. 1).


Origins, distribution and expression of the Duarte-2 (D2) allele of galactose-1-phosphate uridylyltransferase.

Carney AE, Sanders RD, Garza KR, McGaha LA, Bean LJ, Coffee BW, Thomas JW, Cutler DJ, Kurtkaya NL, Fridovich-Keil JL - Hum. Mol. Genet. (2009)

Proposed origins and relationship of the p.N314, D1 and D2 alleles of human GALT. The predicted ancestral human GALT allele carries the (GTCA)3 + p.L218(CTA) + p.D314 sequences found in other hominid species. The p.D314N substitution occurs early in human evolution, and the 4 bp 5′ deletion and p.L218(TTA) silent substitution occur later, on distinct branches of the tree. The three intronic base changes reported to exist in cis with D2 alleles [c.378-27G>C or IVS4-27G > C, c.508-24G > A or IVS5-24G > A and c.507 + 62G > A or IVS5-62G > A (20–22), not illustrated here] presumably occurred subsequent to, or concurrently with, the 4 bp 5′ deletion on the D2 branch of the tree.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2667289&req=5

DDP080F1: Proposed origins and relationship of the p.N314, D1 and D2 alleles of human GALT. The predicted ancestral human GALT allele carries the (GTCA)3 + p.L218(CTA) + p.D314 sequences found in other hominid species. The p.D314N substitution occurs early in human evolution, and the 4 bp 5′ deletion and p.L218(TTA) silent substitution occur later, on distinct branches of the tree. The three intronic base changes reported to exist in cis with D2 alleles [c.378-27G>C or IVS4-27G > C, c.508-24G > A or IVS5-24G > A and c.507 + 62G > A or IVS5-62G > A (20–22), not illustrated here] presumably occurred subsequent to, or concurrently with, the 4 bp 5′ deletion on the D2 branch of the tree.
Mentions: As a first step toward understanding the origin and relationship of the p.N314D (rs2070074 at position 34,639,442 of chromosome 9 in NCBI Build 36.1) and 5′ variations in human GALT, we performed cross-species comparisons of the appropriate coding and non-coding sequences from representative human, non-human hominid (e.g. chimpanzee), non-hominid primate (e.g. macaque) and non-primate placental mammalian (e.g. mouse) species. All species examined, other than human, encoded D rather than N at position 314 (Table 1), strongly implicating D314 as the ancestral GALT allele; the variant that is predominant among modern humans is therefore most appropriately termed p.D314 N (Fig. 1).

Bottom Line: Patients with Duarte galactosemia demonstrate reduced GALT activity and carry one profoundly impaired GALT allele (G) along with a second, partially impaired GALT allele (Duarte-2, D2).The p.N314D substitution, however, is not; it is found together with a silent polymorphism, p.L218(TTA), on functionally normal Duarte-1 alleles (D1, also called Los Angeles or LA alleles).Here we present allele-specific qRT-PCR evidence that D2 alleles express less mRNA in vivo than their wild-type counterparts; the difference is small but statistically significant.

View Article: PubMed Central - PubMed

Affiliation: Emory University, Atlanta, USA

ABSTRACT
Duarte galactosemia is a mild to asymptomatic condition that results from partial impairment of galactose-1-phosphate uridylyltransferase (GALT). Patients with Duarte galactosemia demonstrate reduced GALT activity and carry one profoundly impaired GALT allele (G) along with a second, partially impaired GALT allele (Duarte-2, D2). Molecular studies reveal at least five sequence changes on D2 alleles: a p.N314D missense substitution, three intronic base changes and a 4 bp deletion in the 5' proximal sequence. The four non-coding sequence changes are unique to D2. The p.N314D substitution, however, is not; it is found together with a silent polymorphism, p.L218(TTA), on functionally normal Duarte-1 alleles (D1, also called Los Angeles or LA alleles). The HapMap database reveals that p.N314D is a common human variant, and cross-species comparisons implicate D314 as the ancestral allele. The p.N314D substitution is also functionally neutral in mammalian cell and yeast expression studies. In contrast, the 4 bp 5' deletion characteristic of D2 alleles appears to be functionally impaired in reporter gene transfection studies. Here we present allele-specific qRT-PCR evidence that D2 alleles express less mRNA in vivo than their wild-type counterparts; the difference is small but statistically significant. Furthermore, we characterize the prevalence of the 4 bp deletion in GG, NN and DG populations; the deletion appears exclusive to D2 alleles. Combined, these data strongly implicate the 4 bp 5' deletion as a causal mutation in Duarte galactosemia and suggest that direct tests for this deletion, as proposed here, could enhance or supplant current tests, which define D2 alleles on the basis of the presence and absence of linked coding sequence polymorphisms.

Show MeSH
Related in: MedlinePlus