Functional analysis of the -95G>T PTGDRpromoter variant in asthmatic patients

Resumen

Background:Allergy and asthma arethought to be determined by the interac-tion between several genes and differentenvironmental factors. Among these genesPTGDR has been strongly associated withasthma susceptibility in diverse popula-tions. In the present study we analyze theputative association of the recentlydescribed)95G>T PTGDR SNP withasthma, as well as its implication in puta-tive transcription binding affinity changes.Methods:In this study, 602 Caucasianindividuals were included (351 Asthmaticpatients and 251 non-atopic non-asthmaticcontrols). Asthma was physician-diagnosedand allergy was determined by skin pricktesting.)95G>T Genotype frequencieswere determined by PCR and directSequencing. In silico studies consideringthe)95G>T variation were performedusing ‘‘ElDorado’’ and ‘‘TESS’’ software.Finally EMSA assays were performed withnuclear proteins extracts and the)95G>Tsurrounded region to detect binding affin-ity changes.Results:Nosignificantassociationsbetween the)95G>T SNP and the asthmaor allergy phenotypes were detected. ElDo-rado in silico analysis revealed no differ-ences between the wild type and themutant allele at)95G>T, whereas TESSsoftware predict the binding lost of tran-scription factors GCF and T-Ag when themutant allele is present. However, thesebinding affinity differences were not con-firmed by EMSA assays.Conclusions:A new)95G>T polymor-phism has been recently described by ourgroup in the PTGDR promoter region;however no genetic association wasdetected with asthma or allergy in thisstudy. Although discrepancies weredetected in thein silicoanalysis, we did notdetect binding affinity differences in theEMSA assay what were in agreement withthe ElDoradoin silicoanalysis. This newSNP seems not to have functional implica-tions in the PTGDR promoter activityalthough discrepancies in thein silicomod-els needs to be explained.