Análisis del perfil de microRNA en la eliminación de los progenitores esqueléticos de los espacios interdigitales embrionarios

Résumé

Next-generation sequencing in combination with quantitative polymerase chain reaction analysis revealed a dynamic miRNA signature in the interdigital mesoderm of the chick embryonic limb in the course of interdigit remodelling. MicroRNAs originally discovered in Caenorhabditis elegans, is found in most eukaryotes, including humans; MicroRNA are small, highly conserved non-coding RNA molecules involved in the regulation of gene expression, It is predicted that miRNA account for 1-5% of the human genome and regulate at least 30% of protein-coding genes. Mature miRNA is generated through two-step cleavage of primary miRNA (pri-miRNA), which incorporates into the effector complex RNA-induced silencing complex (RISC). The miRNA functions as a guide by base-pairing with target mRNA to negatively regulate its expression. The level of complementarity between the guide and mRNA target determines which silencing mechanism will be employed; cleavage of target messenger RNA (mRNA) with subsequent degradation or translation inhibition. During this period, 612 previously known chicken miRNAs (gga-miRNAs) and 401 nonidentified sequences were expressed in the interdigital mesoderm. Thirty-six microRNAs, represented by more than 750 reads per million, displayed differential expression between stages HH29 (6 id) and HH32 (7.5 id), which correspond to the onset and the peak of interdigital cell death. Twenty miRNAs were upregulated by at least 1.5-fold, and sixteen were downregulated by at least 0.5-fold. Upregulated miRNAs included miRNAs with recognized proapoptotic functions in other systems (miR-181 family, miR-451 and miR-148a), miRNAs associated with inflammation and cell senescence (miR-21 and miR-146) and miRNAs able to induce changes in the extracellular matrix (miR-30c). In contrast, miRNAs with known antiapoptotic effects in other systems, such as miR-222 and miR-205, became downregulated. In addition, miR-92, an important positive regulator of cell proliferation, was also downregulated. Together, these findings indicate a role for miRNAs in the control of tissue regression and cell death in a characteristic morphogenetic embryonic process based on massive apoptosis.