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Effects of two common polymorphisms in the 3' untranslated regions of estrogen receptor beta on mRNA stability and translatability.

ABSTRACT: BACKGROUND: The present study represents the first attempt to functionally characterize two common single nucleotide polymorphisms (SNPs) in the 3'untranslated regions (3'UTRs) of estrogen receptor beta (ERbeta), focusing on the differences between alleles with regard to mRNA stability and translatability. These two ERbeta SNPs have been investigated for association with disease in a large number of reports. RESULTS: Here we examined allelic expression in breast tumor samples from heterozygous individuals. A significant difference in mRNA levels of the two alleles was observed for one of the SNPs. A cell model system was employed to further investigate potential molecular effects of the two SNPs. We used a modified plasmid, containing the ERbeta promoter and ERbeta 3'UTRs which include the different alleles of investigated SNPs. Quantitative Real-Time PCR was used to determine mRNA levels after inhibition of transcription by actinomycin D, and a luciferase assay was used to determine protein levels. The obtained results suggested that there was no difference in mRNA stability or translatability between the alleles of investigated SNPs. CONCLUSION: Our results indicate that observed associations between ERbeta 3'UTR SNPs and disease susceptibility are due to linkage disequilibrium with another gene variant, rather than the variant itself being the susceptibility factor.

SUBMITTER: Putnik M

PROVIDER: S-EPMC2759954 | biostudies-literature | 2009

REPOSITORIES: biostudies-literature

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Effects of two common polymorphisms in the 3' untranslated regions of estrogen receptor beta on mRNA stability and translatability.

Putnik Milica M Zhao Chunyan C Gustafsson Jan-Ake JA Dahlman-Wright Karin K

BMC genetics 20090915

<h4>Background</h4>The present study represents the first attempt to functionally characterize two common single nucleotide polymorphisms (SNPs) in the 3'untranslated regions (3'UTRs) of estrogen receptor beta (ERbeta), focusing on the differences between alleles with regard to mRNA stability and translatability. These two ERbeta SNPs have been investigated for association with disease in a large number of reports.<h4>Results</h4>Here we examined allelic expression in breast tumor samples from h ...[more]

PMID: 19754929

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Project description:BackgroundA promising application of the huge amounts of genetic data currently available lies in developing a better understanding of complex diseases, such as cancer. Analysis of publicly available databases can help identify potential candidates for genes or mutations specifically related to the cancer phenotype. In spite of their huge potential to affect gene function, no systematic attention has been paid so far to the changes that occur in untranslated regions of mRNA.ResultsIn this study, we used Expressed Sequence Tag (EST) databases as a source for cancer-related sequence polymorphism discovery at the whole-genome level. Using a novel computational procedure, we focused on the identification of untranslated region (UTR)-localized non-coding Single Nucleotide Polymorphisms (UTR-SNPs) significantly associated with the tumoral state. To explore possible relationships between genetic mutation and phenotypic variation, bioinformatic tools were used to predict the potential impact of cancer-associated UTR-SNPs on mRNA secondary structure and UTR regulatory elements. We provide a comprehensive and unbiased description of cancer-associated UTR-SNPs that may be useful to define genotypic markers or to propose polymorphisms that can act to alter gene expression levels. Our results suggest that a fraction of cancer-associated UTR-SNPs may have functional consequences on mRNA stability and/or expression.ConclusionWe have undertaken a comprehensive effort to identify cancer-associated polymorphisms in untranslated regions of mRNA and to characterize putative functional UTR-SNPs. Alteration of translational control can change the expression of genes in tumor cells, causing an increase or decrease in the concentration of specific proteins. Through the description of testable candidates and the experimental validation of a number of UTR-SNPs discovered on the secreted protein acidic and rich in cysteine (SPARC) gene, this report illustrates the utility of a cross-talk between in silico transcriptomics and cancer genetics.

Project description:The serotonin transporter (SERT, SLC6A4) is a Na+-dependent transporter that regulates the availability of serotonin (5-HT, 5-hydroxytryptamine), a key neurotransmitter and hormone in the brain and the intestine. The human SERT gene consists of two alternate promoters that drive expression of an identical SERT protein. However, there are different mRNA transcript variants derived from these two promoters that differ in their 5' untranslated region (5'UTR), which is the region of the mRNA upstream from the protein-coding region. Two of these transcripts contain exon-1a and are abundant in neuronal tissue, whereas the third transcript contains exon-1c and is abundant in the intestine. The 3'UTR is nearly identical among the transcripts. Current studies tested the hypothesis that the UTRs of SERT influence its expression in intestinal epithelial cells (IECs) by controlling mRNA or protein levels. The SERT UTRs were cloned into luciferase reporter plasmids and luciferase mRNA and activity were measured following transient transfection of the UTR constructs into the model IEC Caco-2. Luciferase activity and mRNA abundance were higher than the empty vector for two of the three 5'UTR variants. Calculation of translation index (luciferase activity divided by the relative luciferase mRNA level) revealed that the exon-1a containing 5'UTRs had enhanced translation when compared to the exon-1c containing 5'UTR which exhibited a low translation efficiency. Compared to the empty vector, the SERT 3'UTR markedly decreased luciferase activity. In silico analysis of the SERT 3'UTR revealed many conserved potential miRNA binding sites that may be responsible for this decrease. In conclusion, we have shown that the UTRs of SERT regulate mRNA abundance and protein expression. Delineating the molecular basis by which the UTRs of SERT influence its expression will lead to an increased understanding of post-transcriptional regulation of SERT in GI disorders associated with altered 5-HT availability.

Dataset Information

Effects of two common polymorphisms in the 3' untranslated regions of estrogen receptor beta on mRNA stability and translatability.

Publications

Effects of two common polymorphisms in the 3' untranslated regions of estrogen receptor beta on mRNA stability and translatability.

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