Functional genomics of membrane transporters in human populations

Abstract

Although considerable progress has been made toward characterizing human DNA sequence variation, there remains a deficiency in information on human phenotypic variation at the single-gene level. We systematically analyzed the function of all protein-altering variants of eleven membrane transporters in heterologous expression systems. Coding-region variants were identified by screening DNA from a large sample (n = 247-276) of ethnically diverse subjects. In total, we functionally analyzed 88 protein-altering variants. Fourteen percent of the polymorphic variants (defined as variants with allele frequencies Ն1% in at least one major ethnic group) had no activity or significantly reduced function. Decreased function variants had significantly lower allele frequencies and were more likely to alter evolutionarily conserved amino acid residues. However, variants at evolutionarily conserved positions with approximately normal activity in cellular assays were also at significantly lower allele frequencies, suggesting that some variants with apparently normal activity in biochemical assays may influence occult functions or quantitative degrees of function that are important in human fitness but not measured in these assays. For example, eight (14%) of the 58 variants for which we had measured the transport of at least two substrates showed substrate-specific defects in transport. These variants and the reduced function variants provide plausible candidates for disease susceptibility or variation in clinical drug response.Since the completion of the human genome project, considerable progress has been made in characterizing the nature and degree of human DNA sequence variation (Cargill et al. 1999;Halushka et al. 1999;Patil et al. 2001;Stephens et al. 2001;Leabman et al. 2003). Numerous single nucleotide polymorphisms in all regions of the human genome have been identified, including coding and noncoding regions of genes and large intergenic regions. However, there remains a lack of information connecting genetic polymorphisms to human phenotypic variation. As a consequence, current attempts to predict the functional significance of genetic variants from sequence data alone (from interspecies sequence comparisons or degree of chemical change for amino acid substitutions, as examples) (Sunyaev et al. 2000(Sunyaev et al. , 2001(Sunyaev et al. , 2003Chasman and Adams 2001;Fay et al. 2001;Muller et al. 2001;Henikoff 2001, 2003;Wang and Moult 2001;Ramensky et al. 2002) are lacking in support from empirical data. Furthermore, we have little information on the fraction of natural variants with altered or potentially deleterious function that are harbored in healthy populations. In order to understand variation in normal human physiology and responses to the environment (including, for example, drug-response phenotypes), it is important to understand the range of phenotypic variation that is associated with genetic variation in human populations.Analysis of causal mutations in Mendelian diseases has demonstrated that the majority...

Genetics of P450 oxidoreductase: Sequence variation in 842 individuals of four ethnicities and activities of 15 missense mutations

“…Although an equal number of synonymous and nonsynonymous sites were identified, the nucleotide diversity at various sites ( S ) was considerably greater than that at nonsynonymous sites ( NS ). The ratio of NS / S, used as a measure of selective pressure, was 0.19 for POR, a value much less than one and indicative of selective pressure on the gene (17,18). Selective pressure on POR is comparable with other genes resequenced in large SNP genotyping projects (17,19).…”

Metabolism and Toxicity of Drugs. Two Decades of Progress in Industrial Drug Metabolism

“…The next decade also will bring an appreciation of the functional consequences of genetic polymorphisms in drug transporters (57)(58)(59), in much the same way as has been achieved in the past two decades in the drug-metabolizing enzyme field (22). Developments in the concept of personalized medicine, in which both the selection of a specific therapeutic agent and its associated dosage regimen are tailored to the individual patient on the basis of genetic considerations (60), clearly will have ramifications for drug metabolism in that genetic aspects of each of the processes involved in ADME will need to be better defined than is the case today (61,62).…”

Current Updates in the Genetic Polymorphisms of Human Organic Anion Transporters (OATs)

“…Among the 540 individuals with different ethnicity in these studies, F129L and A399S were only observed in Mexican-American, Q239X and I260R were particularly found in Asian-American, R277W and V281A only occurred in African-American, and A310V was exclusive to European-American [1,59]. The rest of the genetic variants were presented in more than one ethnic group [1,59].…”

“…Later study indicated that Estrone-3-Sulfate (ES) and cimetidine uptake was essentially lost in the R149S, G239X and I260R variants, while R277W and I305F only demonstrated a significantly lower ES uptake comparing to that of the reference transporter. All the other variants assessed in the same study did not show a considerable difference in the uptake of either substrate [1,59]. More recently, Vormfelde et al [56] molecularly investigated two 5' UTR and one non-synonymous SNPs of OAT3, and none of these SNPs was shown to affect the renal clearance of the loop diuretic, torsemide.…”

“…Another non-synonymous SNP was also identified by Xu et al [50] (523A>G, I175V), which was only detected in one Japanese individual (AF=5%). Erdman et al [1] and Urban et al [59] discovered more novel non-silent variants, which attributed to the following amino acid substitutions: F129L, R149S, Q239X, I260R, R277W, V281A, I305F, A310V, A399S and V448I. Among the 540 individuals with different ethnicity in these studies, F129L and A399S were only observed in Mexican-American, Q239X and I260R were particularly found in Asian-American, R277W and V281A only occurred in African-American, and A310V was exclusive to European-American [1,59].…”