| The Wilms’ tumor suppressor gene WT1 is a transcription factor encoding a zinc-finger protein,which plays an important role in the development of mammalian genitourinary system.Mutations of WT1 in mammals give rise to multiple organ defects,including kidney,spleen,and gonads.In zebrafish(Danio rerio),there are two paralogous wt1 genes,naming wt1 a and wt1 b.In this experiment,we examined the roles of both wt1 genes in zebrafish embryonic development by the CRISPR/Cas9 technology.The homozygous wt1 a mutants displayed developmental malformations including pericardial edema,yolk sac edema and failure of glomerulus development while the homozygous wt1 b mutants were phenotypically normal and had no severe phenotypes of nephrogenesis defects that were observed in wt1 b morpholino knockdown embryos.Podocytes are essential elements of the glomerular filtration barrier and relevant to many kidney diseases.To further verify whether there is a relationship between the developmental malformation of homozygous wt1 a mutants and the development of kidney podocytes we selected the podocin and nephrin as the marker genes for podocytes.Loss of wt1 a function disrupted podocyte differentiation and inhibited the expression of podocin and nephrin.However,the expression lever of podocin and nephrin in homozygous wt1 b mutants is similar to normal one.Therefore,wt1 a plays an essential role in the kidney development,whereas wt1 b might not be essential for kidney development.Autophagy is a common and important phenomenon in eukaryotic cells,which plays an important role in maintaining cell homeostasis and cell life activities.Autophagy abnormal is closely related to cell growth and development and various diseases.We found autophagy signal was greatly reduced in wt1 a mutants.To better understand the relationship between autophagy signal and wt1 a function in podocyte development rapamycin was added to treat zebrafish embryos and activate autophagy signal in wt1a-mutated embryos.Then the markers genes’ expression lever were checked.As a result,podocyte injury was ameliorated in wt1 a homozygous mutants by rapamycin treatment.In conclusion,our findings not only confirmed the essential function of wt1 a during kidney development,but also provided an idea for establishing the research model of fish podocyte injury. |
