AMD-associated variants at chromosome 10q26 locus and ARMS2/HTRA1gene expression

Abstract

Purpose: To analyze the relationship between age-related macular degeneration (AMD) associated variants at the chromosome 10q26 locus and ARMS2/HTRA1 expression in human retinas and in vitro systems.

Methods: ARMS2 minigenes with different genotypes at rs2736911 (R38X) and the 3’ UTR indel were constructed. Exogenous ARMS2 transcription from minigenes was assessed in mouse embryonic fibroblasts (MEF) which do not express endogenous ARMS2. Dual luciferase assays were applied to further evaluate the effect of the indel on ARMS2 expression. Genotypes of retinal samples at ARMS2 variants R38X and rs11200638 and HTRA1 variants rs1049331 and rs2293870 were obtained by Taqman assays. Genotypes at the indel of ARMS2 were obtained by PCR and gel assay. RT-PCR and quantitative RT-PCR were applied to measure endogenous ARMS2 or HTRA1 transcripts in human retinal samples (n=82). HTRA1 protein was evaluated by Western blot.

Results: R38X significantly decreases exogenous ARMS2 transcript levels from minigenes in MEF. In contrast, the indel does not significantly change ARMS2 transcript levels . Dual luciferase assays further show that the indel does not influence gene expression. Although R38X appears to decrease the stability of exogenous ARMS2 transcripts in cultured cells, genotypes at R38X, the indel and rs11200638 do not significantly change the level of either ARMS2 or HTRA1 transcripts in human retinas. Furthermore, two synonymous variants rs1049331 and rs2293870 in HTRA1 do not change its protein level either.

Conclusions: AMD-associated variants at chromosome 10q26 locus do not significantly change the gene expression of either ARMS2 or HTRA1 in vivo, suggesting that the influence of these variants on risk of AMD may be through other biological mechanisms.

Keywords: 412 age-related macular degeneration • 533 gene/expression • 539 genetics