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Thomas Sauer, Sabine Kurth, Frank Holz, Tim Krohne; DICER1 is regulated via cAMP in an EPAC/Rap1-dependent manner in human retinal pigment epithelial cells - implications for age-related macular degeneration. Invest. Ophthalmol. Vis. Sci. 2013;54(15):3656.
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DICER1 deficiency has recently been implicated in the pathogenesis of geographic atrophy (GA) due to age-related macular degeneration (AMD). However, little is known about DICER1 gene regulation in RPE cells. Cyclic adenosine monophosphate (cAMP) regulates DICER1 in human melanocytes. Therefore, we investigated the effects of cAMP on DICER1 gene expression in human RPE cells
The human RPE cell line ARPE-19 was treated with adenylate cyclase agonist forskolin, protein kinase A (PKA) inhibitor H89, Rap1 inhibitor GGTI-298, and Epac agonist 8-pCPT-2-0-Me-cAMP, both as single and combination treatments. Subsequently, cells were lysed and total mRNA was isolated. To assess DICER1 gene expression, real-time quantitative PCR was carried out using DICER1- specific primers: forward 5-CCCGGCTGAGAGAACTTACG-, reverse 5-CTGTAACTTCGACCAACACCTTTAAA-3
In forskolin-treated cells, increased cAMP levels resulted in downregulation of DICER1 (0.04 ± 0.036 relative gene expression). Inhibition of PKA did not prevent cAMP-induced downregulation (0.47 ± 0.41) whereas inhibition of Rap1 did (5.67 ± 5.23). Even without forskolin treatment, activation of Epac downregulated DICER1 (0.21 ± 0.21) whereas inhibition of Rap1 resulted in an upregulation (4.37 ± 3.52)
cAMP-mediated downregulation of DICER1 in human RPE cells is not solely explained by PKA action but involves the Epac/Rap1 pathway. As DICER1 has been demonstrated to prevent Alu RNA-mediated damage in RPE, Rap1 represents a novel therapeutic target for advanced dry AMD
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