Effect of cilp on the development of the intermuscular bones and vertebra in zebrafish

Abstract:

To investigate the molecular mechanism of cilp in the development of vertebrae and intermuscular bones in zebrafish, we established a zebrafish cilp^-/- homozygous mutant strain by CRISPR/Cas9. The bones staining of wild type zebrafish and cilp^-/- strain showed that the loss function of cilp caused abnormal fusion of vertebrae and deficiency of neural spine in mutants, and the number of intermuscular bones in cilp^-/- strain was significantly reduced by 10.27%, while the length of intermuscular bones did not change significantly. Furthermore, we analyzed the expression levels of 14 bone development-related genes in five embryonic development stages and 90 dph (days post hatching) adult fish skeletal tissue. The results showed that the col1a1a, sp7, smad4a, and smad5 genes were significantly differentially expression throughout the embryonic development stages compared to the wild type, the expression levels of bmp2a, bmp2b, smad5, sp7, runx2a, runx2b, and bglap in the skeleton of the 90 dph mutant adult fish were significantly lower than those of the wild type zebrafish, which indicated that the cilp gene knockout down-regulated the expression levels of genes in BMPs and SMADs families which consequentially repressed the expression of genes related to osteoblast development. In summary, dysfunctions of the cilp gene might affect the development of intermuscular bones and skeleton in zebrafish by suppressing the BMPs signaling pathway to affect the development of osteoblasts and bones formation.