p-SMAD2/3 and DICER promote pre-miR-21 processing during pressure overload-associated myocardial remodeling. p-SMAD2/3 and DICER promote pre-miR-21 processing during pressure overload-associated myocardial remodeling. García R, Nistal JF, Merino D, Price NL, Fernández-Hernando C, Beaumont J, González A, Hurlé MA, Villar AV. Biochim Biophys Acta. 2015 Jul;1852(7):1520-30. doi: 10.1016/j.bbadis.2015.04.006.2015-07-11T22:00:00Z<div style="text-align:justify;"><br></div><p style="text-align:justify;"><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"><span class="ms-rteThemeForeColor-2-5 ms-rteThemeFontFace-1 ms-rteFontSize-2" style="font-weight:bold;">Abstract</span><br></span></p><div style="color:#000000;font-family:arial, helvetica, clean, sans-serif;text-align:justify;"><p style="margin-bottom:0.5em;font-size:1.04em;"><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2">Transforming growth factor-β (TGF-β) induces miR-21 expression which contributes to fibrotic events in the left ventricle (LV) under </span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">pressure</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> overload. SMAD effectors of TGF-β signaling interact with DROSHA to </span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">promote</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> primary miR-21 </span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">processing</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> into precursor miR-21 (</span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">pre-miR-21</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2">). We hypothesize that p-SMAD-2 and -</span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">3</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> also interact with DICER1 to regulate the </span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">processing</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> of </span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">pre-miR-21</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> to mature miR-21 in cardiac fibroblasts under experimental and clinical </span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">pressure</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> overload. The subjects of the study were mice undergoing transverse aortic constriction (TAC) and patients with aortic stenosis (AS). In vitro, NIH-3T3 fibroblasts transfected with </span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">pre-miR-21</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2">responded to TGF-β1 stimulation by overexpressing miR-21. Overexpression and silencing of SMAD2/</span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">3</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> resulted in higher and lower production of mature miR-21, respectively. DICER1 co-precipitated along with SMAD2/</span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">3</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> and both proteins were up-regulated in the LV from TAC-mice. </span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">Pre-miR-21</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> was isolated bound to the DICER1 maturation complex. Immunofluorescence analysis revealed co-localization of </span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">p-SMAD2</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2">/</span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">3</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> and DICER1 in NIH-3T3 and mouse cardiac fibroblasts. DICER1-</span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">p-SMAD2</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2">/</span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">3</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> protein-protein interaction was confirmed by in situ proximity ligation assay. </span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">Myocardial</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> up-regulation of DICER1 constituted a response to </span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">pressure</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> overload in TAC-mice. </span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">DICER</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> mRNA levels correlated directly with those of TGF-β1, SMAD2 and SMAD3. In the LV from AS patients, </span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">DICER</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> mRNA was up-regulated and its transcript levels correlated directly with TGF-β1, SMAD2, and SMAD3. Our results support that </span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">p-SMAD2</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2">/</span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">3</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2">interacts with DICER1 to </span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">promote</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> </span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">pre-miR-21</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> </span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">processing</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> to mature miR-21. This new TGFβ-dependent regulatory mechanism is involved in miR-21 overexpression in cultured fibroblasts, and in the </span><span class="highlight ms-rteThemeFontFace-1 ms-rteFontSize-2">pressure</span><span class="ms-rteThemeFontFace-1 ms-rteFontSize-2"> overloaded LV of mice and human patients.</span></p></div><p><a href="https://www.ncbi.nlm.nih.gov/pubmed/25887159">Biochim Biophys Acta</a>. 2015 Jul;1852(7):1520-30. doi: 10.1016/j.bbadis.2015.04.006. Epub 2015 Apr 15.<br></p>128