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3’-RNA Adenylation Enzymes Control Allergic Inflammation Through Regulation of MicroRNA Degradation
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A1197 - 3’-RNA Adenylation Enzymes Control Allergic Inflammation Through Regulation of MicroRNA Degradation
Author Block: A. Korde1, S. Mehta2, S. Takyar3; 1Internal Medicine, Yale school of Medicine, New Haven, CT, United States, 2Genetics, Yale school of Medicine, New Haven, CT, United States, 3Pulm and Crit Care, Yale School of Med, New Haven, CT, United States.
Introduction: We have previously shown that miR-1 downregulation plays a critical role in allergic inflammation. This regulation is mediated through VEGF signaling and involves the degradation of mature miRNA. 3' end modification of RNAs is one of the most common mechanisms triggering miRNA degradation.
Hypothesis: 3'end RNA modification enzymes regulate miRNA levels and control the severity of allergic inflammation.
Methods: 3' modifications of the microRNAS were identified by high depth small RNA-sequencing. Lung-specific VEGF or IL13 overexpression were induced by adding Doxycycline to the drinking water of CC10-VEGF or IL-13 transgenic mice. Airway allergic inflammation was induced by sensitization and challenge of C57/BL6 mice with house dust mute (HDM). Levels of miR-1 and its precursors were measured by quantitative real time RT-PCR. We created knock down vectors by cloning shRNA against RNA-modifying enzymes in a lentiviral backbone. These vectors were delivered intranasally to various models.
Results: VEGF stimulation increased the prevalence of 3' adenylated miR-1 isoforms. These modifications were accompanied by an upregulation of the 3' RNA adenylation enzymes, poly A RNA polymerase D4 and D5 (PAPD4 and 5), compared to other 3' modification enzymes. PAPD upregulation immediately preceded miR-1 downregulation and was necessary for its degradation. Furthermore, knockdown of PAPDs increased miR-1 levels in endothelial cells. We assessed the pathologic significance of PAPDs by testing the effect of their knockdown in VEGF transgenic, IL-13 transgenic and HDM-induced asthma models. PAPD knockdown prevented miR-1 degradation in all of these models and significantly reduced lung inflammation. In tissue biopsies from chronic rhinosinusitis (CRS) patients, the levels of PAPD4 were higher in the eosinophilic (allergic) tissue samples and correlated with miR-1 levels.
Conclusion: 3' RNA adenylation enzymes control miRNA levels by triggering their degradation and are necessary for the propagation of allergic airway inflammation. These enzymes can be specifically targeted for diagnostic and therapeutic purposes
Author Block: A. Korde1, S. Mehta2, S. Takyar3; 1Internal Medicine, Yale school of Medicine, New Haven, CT, United States, 2Genetics, Yale school of Medicine, New Haven, CT, United States, 3Pulm and Crit Care, Yale School of Med, New Haven, CT, United States.
Introduction: We have previously shown that miR-1 downregulation plays a critical role in allergic inflammation. This regulation is mediated through VEGF signaling and involves the degradation of mature miRNA. 3' end modification of RNAs is one of the most common mechanisms triggering miRNA degradation.
Hypothesis: 3'end RNA modification enzymes regulate miRNA levels and control the severity of allergic inflammation.
Methods: 3' modifications of the microRNAS were identified by high depth small RNA-sequencing. Lung-specific VEGF or IL13 overexpression were induced by adding Doxycycline to the drinking water of CC10-VEGF or IL-13 transgenic mice. Airway allergic inflammation was induced by sensitization and challenge of C57/BL6 mice with house dust mute (HDM). Levels of miR-1 and its precursors were measured by quantitative real time RT-PCR. We created knock down vectors by cloning shRNA against RNA-modifying enzymes in a lentiviral backbone. These vectors were delivered intranasally to various models.
Results: VEGF stimulation increased the prevalence of 3' adenylated miR-1 isoforms. These modifications were accompanied by an upregulation of the 3' RNA adenylation enzymes, poly A RNA polymerase D4 and D5 (PAPD4 and 5), compared to other 3' modification enzymes. PAPD upregulation immediately preceded miR-1 downregulation and was necessary for its degradation. Furthermore, knockdown of PAPDs increased miR-1 levels in endothelial cells. We assessed the pathologic significance of PAPDs by testing the effect of their knockdown in VEGF transgenic, IL-13 transgenic and HDM-induced asthma models. PAPD knockdown prevented miR-1 degradation in all of these models and significantly reduced lung inflammation. In tissue biopsies from chronic rhinosinusitis (CRS) patients, the levels of PAPD4 were higher in the eosinophilic (allergic) tissue samples and correlated with miR-1 levels.
Conclusion: 3' RNA adenylation enzymes control miRNA levels by triggering their degradation and are necessary for the propagation of allergic airway inflammation. These enzymes can be specifically targeted for diagnostic and therapeutic purposes
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