Indiana University-Purdue University Indianapolis (IUPUI) / Indoleamine 2,3-dioxygenase (IDO) is a rate limiting enzyme in tryptophan
catabolism that has been implicated in the pathogenesis of a number of diseases.
Large interindividual variability in IDO activity in the absence of stimuli and as the
result of therapy induced changes has been reported. This variability has the potential
to contribute to susceptibility to disease and to interindividual variability in
therapeutic response.
To identify genetic variations that might contribute to interindividual
variability in IDO activity, we resequenced the exons, intron/exon borders and 1.3 kb
of the 5’-flanking region of the INDO gene in 48 African-American (AA) and 48
Caucasian-American (CA) subjects from the Coriell DNA Repository. A total of 24
INDO variants were identified. Seventeen of these were in exons, introns, or
exon/intron boundries, while seven were within 1.3 kb upstream of the translation
start site. Seventeen are novel and 7 were previously identified.
When transiently expressed in COS-7 or HEK293 cells the amino acid
sequence change in Arg77His resulted in significant decrease in activity, and it
reduced the Vmax of IDO. The Arg77His variant and the 9 bp deletion resulted in
nearly complete loss of enzyme activity and a lack of detectable protein expression.
The function of the Arg77His variant IDO was restored in a dose dependent
manner by the heme analog hemin; but there was no associated increase in IDO
protein. Cellular heme concentration was higher in cells transfected with the wild
type and Ala4Thr variant constructs, but not in cells transfected with the Arg77His
variant. The heme synthesis inhibitor, succinylacetone (SA), blocked IDO activity in
cells transfected with Arg77His.
We identified 22 putative transcription binding sites within the 1.3 kb
upstream of the translation start site. Two of the SNPs were located in GATA3 and
FOXC1 sites. A specific 3-SNP haplotype reduced promoter activity when transiently
transfected into 2 different cell lines.
We conclude that there are naturally occurring genetic variants in the INDO
gene which affect both expression and activity. These results make clear that
interindividual variability in IDO activity at baseline or in response to therapy may be
in part due to inherited genetic variability.
| Identifer | oai:union.ndltd.org:IUPUI/oai:scholarworks.iupui.edu:1805/1704 |
| Date | 13 October 2008 |
| Creators | Arefayene, Million |
| Contributors | Flockhart, David A., Vasko, Michael R., Queener, Sherry F., Safa, Ahmad R., Wilkes, David S. |
| Source Sets | Indiana University-Purdue University Indianapolis |
| Language | en_US |
| Detected Language | English |
| Type | Thesis |
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