Investigation of polymorphisms in the promoter region of the gulo gene in dogs and their effects on promoter activity

L-ascorbic acid, originally called Vitamin C, is an important vitamin for the health of mammals. The enzyme that catalyzes the terminal step in the biosynthesis of vitamin C is encoded by a gene called GULO. While GULO gene has become a pseudogene in humans, hominoid primates, bony fishes, perching birds (passeriformes order), guinea-pigs and some of the fruit bats, it is still functional in all other mammals including dogs. The aim of the study is to detect the polymorphisms and promoter motifs in promoter region of the GULO gene in dogs and prove the effect of these polymorphisms on gene activity. For this purpose, genomic DNAs were isolated from blood samples collected from 50 pariah dogs. This region was amplified by the PCR method with the primers designed and sequenced for the GULO gene promoter region. Polymorphisms have been detected on the sequences that were aligned by Mega7 software and promoter motifs have also been searched within the amplified region. In addition to the investigation of polymorphisms in promoter regions, the splicing motifs of exon-intron junction region (intron 1 and 3 are joined with exon 2) of GULO gene and polymorphisms in this region were determined in 10 dogs. Upon analysis of results, polymorphism in 9 different positions and 10 promoter motifs in the promoter region were detected. Furthermore, 3 splicing motifs and 6 single nucleotide polymorphisms were detected in exon-intron junction region of GULO gene. One of the polymorphisms detected in the promoter region has been found to be located on the common nucleotides of two overlapping promoter motifs. A potential nucleotide change in these motifs may affect the transcription process, as it may affect the binding of motif-specific regulatory elements such as transcription factors and RNA Pol II. Similarly, single nucleotide polymorphisms which are found in promoter region but not in motifs may also effect the transcription level. Since two of the six detected polymorphisms in the exon-intron junction region are located on splicing motifs, the splicing mechanism of the mRNA of GULO gene may be affected.