Abstract
BackgroundOver 4 million single nucleotide polymorphisms (SNPs) are currently reported to exist within the human genome. Only a small fraction of these SNPs alter gene function or expression, and therefore might be associated with a cell phenotype. These functional SNPs are consequently important in understanding human health. Information related to functional SNPs in candidate disease genes is critical for cost effective genetic association studies, which attempt to understand the genetics of complex diseases like diabetes, Alzheimer's, etc. Robust methods for the identification of functional SNPs are therefore crucial. We report one such experimental approach.ResultsSequence conserved between mouse and human genomes, within 5 kilobases of the 5-prime end of 176 GPCR genes, were screened for SNPs. Sequences flanking these SNPs were scored for transcription factor binding sites. Allelic pairs resulting in a significant score difference were predicted to influence the binding of transcription factors (TFs). Ten such SNPs were selected for mobility shift assays (EMSA), resulting in 7 of them exhibiting a reproducible shift. The full-length promoter regions with 4 of the 7 SNPs were cloned in a Luciferase based plasmid reporter system. Two out of the 4 SNPs exhibited differential promoter activity in several human cell lines.ConclusionsWe propose a method for effective selection of functional, regulatory SNPs that are located in evolutionary conserved 5-prime flanking regions (5'-FR) regions of human genes and influence the activity of the transcriptional regulatory region. Some SNPs behave differently in different cell types.
Highlights
Over 4 million single nucleotide polymorphisms (SNPs) are currently reported to exist within the human genome
Among the group of Single Nucleotide Polymorphism (SNP) located within coding regions of genes and causing a change in the peptide sequence or among SNPs located within promoters, a majority may not influence the overall activity of the protein or the gene expression
From a total of approximately 800 SNPs in proximal 5 kb regions, less than 200 were mapped to regions of mouse-human genome conservation. Of these approximately 200 SNPs, 36 were predicted to influence Transcription Factor (TF) binding, in regions of sequence conservation of over 70% in human-mouse; the alignments for two such regions are indicated in additional files [See Additional file 2]
Summary
Over 4 million single nucleotide polymorphisms (SNPs) are currently reported to exist within the human genome. A small fraction of these SNPs alter gene function or expression, and might be associated with a cell phenotype. These functional SNPs are important in understanding human health. Information related to functional SNPs in candidate disease genes is critical for cost effective genetic association studies, which attempt to understand the genetics of complex diseases like diabetes, Alzheimer's, etc. SNPs are the focus of a large number of human genetics studies attempting to understand their impact on complex diseases like Alzheimers, Parkinsons, diabetes, etc. Fewer attempts have been made to predict and validate functional promoter SNPs [8]
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