Requirement of miR-145 in zebrafish neural crest development.

STEEMAN, TOMÁS J.; CALCATERRA, NORA B.; WEINER, ANDREA M.J.

Buenos Aires

Congreso; LASDB Meeting 2019; 2019

The neural crest is a transient and multipotent cell population that gives rise to a diverse cell lineage. A complex gene regulatory network controlling the specification, delamination, migration, and differentiation of thiscell type has been thoroughly described. However, the role of post-transcriptional factors, such as miRNAs, has not been deeply characterized yet.Sox9 is required for determination of the chondrogenic cell lineage in the cranial neural crest. The expression of Sox9 is regulated by miR-145 during chondrogenic differentiation in human and mouse cultured cells, but scant information exists regarding the role of this miRNA during embryogenesis. Metformin treatment of cultured cells caused the overexpression of miR-145, as well as aberrant expression of neural crest markers in zebrafish embryos. The goal of this work was to assess the function of miR-145 during the neural crest development in zebrafish. In-silico analysis of sox9b 3?UTR showed the presence of one miR-145 binding site. Furthermore, the analysis of miR-145 promoter revealed the presence of mul- tiple putative binding sites for SOX9, suggesting a regulatory feedback loop. Overexpression of miR-145 or metformin treatment in zebrafish embryos caused aberrant craniofacial development, low melanin levels, and elevated apoptosis throughout the embryo. Besides, the expression of sox9b, sox10, and dct was analized by RT-qPCR and whole-mount in situ hybridization. Using CRISPR/Cas9 technology, two different knock-out miR-145 mutant lines were generated to further assess the role of miR-145 in the gene regulatory network governing neural crest development.