Genetic variants modulate the PTGDR expression
- García-Sánchez, A 12
- Marcos-Vadillo, E 13
- Isidoro-García, M 123
- Sanz, C 12
- Moreno-Rodilla, E 13
- Lorente, F 123
- Dávila, I 123
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Instituto de Investigación Biomédica de Salamanca
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2
Universidad de Salamanca
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Hospital Universitario de Salamanca
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ISSN: 0105-4538, 1398-9995
Year of publication: 2016
Pages: 598-598
Congress: European Academy of Allergy and Clinical Immunology Congress
Type: Conference paper
Abstract
Background:Asthma and allergy are com-plex diseases that result from interactionbetween the genetic background and envi-ronmental factors. PTGDR, a receptor ofPGD2 (Prostaglandin D2) has beenreported as a candidate gene in asthma.The goal of this study was to determinethe contribution of different promoter hap-lotype variants toPTGDRgene expressionin cell culture.Method:The A549 cell line was trans-fected with fourPTGDRexpression plas-mids. Wild-typePTGDRcDNA clonedinto a pCMV6-entry vector was the originof thePTGDRconstructs. Using theXhoIandBglII restriction enzymes we replacedthe constitutive cytomegalovirus promoterof pCMV6-PTGDRby one of the fourpromoter sequences ofPTGDRwith thefollowingalternativehomozygous haplotypes:CTCT, CCCT, TCCT andCCCC (positions613,549,441 and197). After 24 h of transient transfection,cells were collected and total RNA wasisolated. Relative quantitative PCR ofPTGDRwas performed using SYBRGreen I Master in a Light Cycler 480instrument. Fold induction was calculatedusing the comparative Ct method. TheGAPDHgene was used as internal control.Results:We found significant differencesin thePTGDRexpression among the hap-lotype variants after transient transfectionanalysis in the A549 cell line. The haplo-type with the CCCC sequence (613C,549C,441C, and197C) showed thehighestPTGDRexpression, while theTCCT haplotype showed a lowest expres-sion.PTGDRexpression after normaliza-tion (meanSD) was expressed as foldincrements, considering the wild variantCTCT as value 1: CCCC: 1.400.08;CCCT: 1.040.12; TCCT: 0.520.04,(P<0.039) for CCCC and TCCT pairwisecomparison by analysis of variance.Conclusion:Our results suggest a differen-tialPTGDRexpression according to thepromoter haplotype combination. In previ-ous reports the CCCC genetic variant alsoexhibit a high promoter activity in reportergene assays and was associated to predis-position to asthma