Evaluation of aldehyde dehydrogenase 1 promoter polymorphisms identified in human populations - PubMed (original) (raw)

Comparative Study

Evaluation of aldehyde dehydrogenase 1 promoter polymorphisms identified in human populations

John P Spence et al. Alcohol Clin Exp Res. 2003 Sep.

Abstract

Background: Cytosolic aldehyde dehydrogenase, or ALDH1A1, functions in ethanol detoxification, metabolism of neurotransmitters, and synthesis of retinoic acid. Because the promoter region of a gene can influence gene expression, the ALDH1A1 promoter regions were studied to identify polymorphism, to assess their functional significance, and to determine whether they were associated with a risk for developing alcoholism.

Methods: Sequence analysis was performed in the promoter region by using Asian, Caucasian, and African American subjects. The resulting polymorphisms were assessed for frequency in Asian, Caucasian, Jewish, and African American populations and tested for associations with alcohol dependence in Asian and African American populations of alcoholics and controls. The functional significance of each polymorphism was determined through in vitro expression analysis by using HeLa and HepG2 cells.

Results: Two polymorphisms, a 17 base pair (bp) deletion (-416/-432) and a 3 bp insertion (-524), were discovered in the ALDH1A1 promoter region: ALDH1A1*2 and ALDH1A1*3, respectively. ALDH1A1*2 was observed at frequencies of 0.035, 0.023, 0.023, and 0.012 in the Asian, Caucasian, Jewish, and African American populations, respectively. ALDH1A1*3 was observed only in the African American population, at a frequency of 0.029. By using HeLa and HepG2 cells for in vitro expression, the activity of the luciferase reporter gene was significantly decreased after transient transfection of ALDH1A1*3-luciferase compared with the wild-type construct ALDH1A1*1-luciferase. In an African American population, a trend for higher frequencies of the ALDH1A1*2 and ALDH1A1*3 alleles was observed in a population of alcoholics (p = 0.03 and f = 0.12, respectively) compared with the control population.

Conclusions: ALDH1A1*2 and ALDH1A1*3 may influence ALDH1A1 gene expression. Both ALDH1A1*2 and ALDH1A1*3 produce a trend in an African American population that may be indicative of an association with alcoholism; however, more samples are required to validate this observation. The underlying mechanisms contributing to these trends are still unknown.

PubMed Disclaimer

Figures

Fig. 1

Fig. 1

Sequence analysis of polymorphism in the human ALDH1A1 promoter region. (A) ALDH1A1*2: the box highlights the 17-bp sequence (−416/−432) that was deleted from ALDH1A1*1 to generate ALDH1A1*2. (B) ALDH1A1*3: the box designates the 3-bp insertion (−524). Arrows indicate common flanking nucleotides, and G, A, T, and C represent guanine, adenine, thiamine, and cytosine, respectively.

Fig. 2

Fig. 2

The human ALDH1A1 promoter region. Sequence analysis was performed on the region spanning −699 to +139 with respect to the transcriptional start point (+1). The magnified region extending from −586 to −372 designates the fragment that was amplified during the genotyping assay. Both polymorphisms are distinguished in this diagram: the 3-bp insertion (−524) and the 17-bp deletion (−416/−432). (A) The deletion of five possible fragments results in the same ALDH1A1*2 sequence due to flanking 5′-GGTC-3′ repeats. (B) Three possible insertions at −524, −523, or −519 generate the same ALDH1A1*3 sequence.

Fig. 3

Fig. 3

Autoradiogram of ALDH1A1*1, ALDH1A1*2, and ALDH1A1*3 genotypes. PCR was used to generate [_α_-33P]deoxycytidine triphosphate-radiolabeled fragments that were later separated on a 6% acrylamide gel by using electrophoresis. Each allele, ALDH1A1*1, ALDH1A1*2, and ALDH1A1*3, can be readily identified with this procedure. (A) A homozygous Caucasian with the genotype ALDH1A1*1/*1, the wild-type genotype. (B) A homozygous Asian with the genotype ALDHlA1*2/*2. (C) A heterozygous African American with the genotype ALDH1A1*1/*3.

Fig. 4

Fig. 4

Functional importance of ALDH1A1 promoter polymorphisms. Plasmids ALDH1A1*1, ALDH1A1*2, and ALDH1A1*3 were transfected into HepG2 and HeLa cells; luc+ denotes the luciferase gene. Each respective fragment amplified from the ALDH1A1 promoter extended from −659 to +39 with respect to the transcriptional start point, and the polymorphisms, the deletion (−416/−432) and the insertion (−524), are noted. The activity of each construct was normalized by co-transfection of the internal control plasmid, pRL-CMV, and was expressed as fold change compared with the activity of ALDH1A1*1-luc. The mean ± SEM are representative of the results from three to six independent transfections performed in triplicate with two different plasmid preparations. The significance of the difference between mean values within and between multiple constructs was analyzed with ANOVA. ALDH1A1*3-luc significantly decreased luciferase expression in both Hep G2 and HeLa cells compared with ALDH1A1*1-luc (*p < 0.05). No significant change in expression was associated with ALDH1A1*2-luc.

Similar articles

Cited by

References

    1. Ambroziak W, Izaguirre G, Pietruszko R. Metabolism of retinaldehyde and other aldehydes in soluble extracts of human liver and kidney. J Biol Chem. 1999;274:33366–33373. - PubMed
    1. Brien JF, Loomis CW. Pharmacology of acetaldehyde. Can J Physiol Pharmacol. 1983;61:1–22. - PubMed
    1. Carr LG, Foroud T, Stewart T, Castelluccio P, Edenberg HJ, Li T-K. Influence of ADH1B polymorphism on alcohol use and its subjective effects in a Jewish population. Am J Med. 2002;112:138–143. - PubMed
    1. Chan AW. Racial differences in alcohol sensitivity. Alcohol Alcohol. 1986;21:93–104. - PubMed
    1. Crabb DW, Pinairs J, Hasanadka R, Fang M, Leo MA, Lieber CS, Tsukamoto H, Motomura K, Miyahara T, Ohata M, Bosron W, Sanghani S, Kedishvili N, Shiraishi H, Yokoyama H, Miyagi M, Ishii H, Bergheim I, Menzl I, Parlesak A, Bode C. Alcohol and retinoids. Alcohol Clin Exp Res. 2001;25(Suppl 5):207S–217S. - PubMed

Publication types

MeSH terms

Substances

Grants and funding

LinkOut - more resources