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Inas F Aboobakar, Xue Qin, W Daniel Stamer, R Rand Allingham, Michael A Hauser; A lncRNA in the LOXL1 locus modulates expression of genes relevant to exfoliation glaucoma pathobiology. Invest. Ophthalmol. Vis. Sci. 2016;57(12):790. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Genetic variants in the LOXL1 locus are strongly associated with risk for developing exfoliation glaucoma (XFG) in all populations studied to date. We recently demonstrated that these variants alter promoter activity for the long non-coding RNA (lncRNA) LOXL1-AS1 (Hauser et al, HMG 2015). However, the functional mechanism whereby altered expression of this lncRNA leads to the XFG disease phenotype remains unknown. Given that lncRNAs have important regulatory roles at both neighboring and distant loci, the goal of this project was to investigate global gene expression changes in response to altered LOXL1-AS1 expression.
Immortalized human lens epithelial cells (HLE-B3) were transfected with an siRNA targeting either the LOXL1-AS1 lncRNA or a negative control siRNA. Each condition was tested in triplicate. Total RNA was extracted from cells after a period of 48 hours, labeled and hybridized to Illumina HumanHT-12 Expression BeadChip arrays containing coverage for approximately 47,000 transcripts. Differential gene expression analysis was performed using the limma package in Bioconductor.
siRNA knockdown led to a 5-fold reduction in expression of LOXL1-AS1 (p<0.01). 109 genes were differentially expressed in response to LOXL1-AS1 knockdown (p<0.05). 73 genes had significantly reduced expression, including the extracellular matrix (ECM) constituents LOXL4, COL6A3, TIMP3, ACTA2, SCG2, PDGFC, LUM, and TNFSF10. Expression of 36 genes was significantly increased in response to LOXL1-AS1 knockdown, including HMOX1 and EFHD2.
The lncRNA LOXL1-AS1 modulates expression of genes involved in collagen fibril formation (COL6A3, LOXL4), ECM degradation (TIMP3), cytoskeleton integrity (ACTA2), calcium ion binding (EFHD2), and response to oxidative stress (HMOX1), all of which play a role in XFG pathophysiology. We previously demonstrated that genetic risk variants alter expression levels of the LOXL1-AS1 lncRNA. The data presented here support the hypothesis that altered LOXL1-AS1 levels disrupt ECM homeostasis by affecting gene expression at multiple loci genome-wide, thereby contributing to the pathogenesis of this common, blinding disease.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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