Posttranscriptional Regulation of LOXL1 Expression Via Alternative Splicing and Nonsense-Mediated mRNA Decay as an Adaptive Stress Response

Berner D, Zenkel M, Pasutto F, Hoja U, Liravi P, Gusek-Schneider GC, Kruse F, Schödel J, Reis A, Schlötzer-Schrehardt U (2017)


Publication Type: Journal article

Publication year: 2017

Journal

Book Volume: 58

Pages Range: 5930-5940

Journal Issue: 13

DOI: 10.1167/iovs.17-22963

Abstract

Alternative mRNA splicing coupled to nonsense-mediated decay (NMD) is a common mRNA surveillance pathway also known to dynamically modulate gene expression in response to cellular stress. Here, we investigated the involvement of this pathway in the regulation of lysyl oxidase-like 1 (LOXL1) expression in response to pseudoexfoliation (PEX)-associated pathophysiologic factors.Transcript levels of LOXL1 isoforms were determined in ocular tissues obtained from donor eyes without and with PEX syndrome. Pseudoexfoliation-relevant cell types, including human Tenon's capsule fibroblasts (hTCF) and trabecular meshwork cells (hTMC), were exposed to puromycin, caffeine, TGF-?1, homocysteine, IL-6, retinoic acid, UV-B radiation, oxidative stress, and mechanical stress for up to 48 hours. Western blot analysis was carried out using antibodies against LOXL1, (phosphorylated-) eukaryotic initiation factor 2-? (eIF2-?), and regulator of nonsense transcripts 2 (UPF2). RNA interference was used to knockdown UPF1-3 and Serine/threonine-protein kinase (SMG1).Constitutive expression of wild-type LOXL1 and alternatively spliced LOXL1-a transcripts was detected in all ocular tissues showing highest levels in trabecular meshwork and differential expression between PEX and control specimens. LOXL1-a transcripts were upregulated in hTCF and hTMC by NMD inhibitors puromycin and caffeine (>=6-fold; P < 0.01) or after knockdown of NMD core factors (>=2-fold; P < 0.05), whereas mRNA and protein levels of LOXL1 were reduced (<=0.8 fold; P < 0.05). Exposure of cells to various PEX-associated (stress) factors, including TGF-?1, UV-B light, oxidative stress, mechanical stress, and retinoic acid enhanced LOXL1-a transcript levels (>=1.5-fold; P < 0.05), while partially downregulating LOXL1 levels (<=0.7-fold; P < 0.05). Stress-induced inhibition of NMD was dependent on phosphorylation of eIF2?.These findings provide evidence for a functional role of alternative splicing coupled to NMD in the posttranscriptional regulation of LOXL1 gene expression and suggest this mechanism to represent a dynamic mode of adapting LOXL1 expression to PEX-associated environmental and nutritional cues.

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APA:

Berner, D., Zenkel, M., Pasutto, F., Hoja, U., Liravi, P., Gusek-Schneider, G.C.,... Schlötzer-Schrehardt, U. (2017). Posttranscriptional Regulation of LOXL1 Expression Via Alternative Splicing and Nonsense-Mediated mRNA Decay as an Adaptive Stress Response. Investigative Ophthalmology & Visual Science, 58(13), 5930-5940. https://dx.doi.org/10.1167/iovs.17-22963

MLA:

Berner, Daniel, et al. "Posttranscriptional Regulation of LOXL1 Expression Via Alternative Splicing and Nonsense-Mediated mRNA Decay as an Adaptive Stress Response." Investigative Ophthalmology & Visual Science 58.13 (2017): 5930-5940.

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