| Budget Amount *help |
¥3,300,000 (Direct Cost: ¥3,300,000)
Fiscal Year 2004: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 2003: ¥1,800,000 (Direct Cost: ¥1,800,000)
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| Research Abstract |
Dioxins alter the expression levels of downstream target genes and affect development and differentiation through the aryl hydrocarbon receptor. However, the spectrum of the dioxin-responsive genes is yet to be delineated. The purpose of the present study is to define the spectrum of dioxin-responsive genes in a comprehensive manner using two complementary approaches : bioinformatic analysis and micro-array analysis.
First, we mapped the xenobiotics responsive elements[XREs] within the putative promoter regions[PPRs] in a semi genome-wide manner. XREs are the consensus transcriptional regulation sequence "gcgtg" which are present in the PPRs of several genes induced by dioxins. Human analysis Sequences 10kb upstream from the transcription start sites[TSSs] of 2,773 genes were obtained by aligning 8,060 cDNA sequences in the human full-length cDNA database of The Institute of Medical Science with the human genome sequences 2,843 out of 2,773 genes possessed at least three XRE consensus s
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equences(gcgtg) within 10kb upstream of the TSS. Total number of XREs within the PPR was 26,787 among the 2,773 genes. The position of the 26,787 XREs from the TSS was ploteted on Fig. 2a. The XRE tended to be present closer to the TSS. The position of the smallest duster among all the XRE dusters within each 10kb upstream sequence tended to be present near the TSS. Mouse analysis : Sequences 10kb upstream from the TSSs of 18,541 genes were obtained by aligning 60,770 cDNA sequences in the FANTOM2 mouse full-length cDNA database of with the mouse genome sequences. 9,762 out of 18,541 genes possessed at least three XRE consensus sequences within 10kb upstream of the TSS. Total number of XREs within the PPR was 74,725 among the 9,762 genes. Again, in mice, the XRE tended to be present closer to the TSS. The position of the smallest cluster among all the XRE clusters within each 10kb upstream sequence trended to be present near the TSS.
Second, we evaluated the expression profile of the mouse embryonic brain exposed to the prototypic dioxin, 2,3,7,8-tetrarchlorodibenzo-p-dioxin (TCDD), and compared with the expression profile of the unexposed brain. The expression values for the 6 hybridizations(3 from experimental group and 3 from the control group) were background-subtracted and normalized. We inferred genes that differentially expressed under TCDD exposure using a robust data-driven method. About 0.37%(46/12488) of the assessed genes showed significant(p<0.0001) change, either increase or decrease, in gene expression(Table 1). Among the 40 genes, 22 genes increased and 24 genes decreased their expression in TCDD-exposed pup brain. Cyp1b1 and Cyp1a1, both of which have been shown previously to be induced by dioxin exposure in vitro, were induced in the list. The changes in the expression level were plotted against the average intensity and the Bonferroni-style gene by gene tests were performed to determine the critical interval. Some of genes down-regulated by dioxin exposure are disease-causing genes : Mid1, Pitx2, and Gata3. Mutations in those genes lead to Opitz syndrome, Rieger syndrome, and HDR syndrome, respectively. Mutation in human MID1 gene is known to cause cleft palate, a cardinal feature of prenatal dioxin exposure in mice. In addition, Glyoxalase 1 (GLO) was down-regulated in the exposed group. A reduction in GLO1 enzyme activity has been demonstrated to result in the accumulation of methylglyoxal, which may be toxic to the developing brain Furthermore, an association study revealed that single nucleotide polymorphism in glyoxalase I as autism susceptibility factor (Junaid et al., 2004). In view of a potential role of xenobiotic toxins in the development of autistic spectrum (Edelson and Cantor, 1998), the biological relevance of GLO1 reduction in the exposed group needs to be further explored. Less
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