RNA-seq for gata1+ cells in add1 mutant zebrafish embryos and siblings at 22 hpf

Adducin 1 (Add1) functions primarily as a membrane cytoskeletal protein, whereas Add1 contains a bipartite nuclear localization signal, implying its special nuclear function. However, the nuclear functional roles of Add1 apart from maintaining cytoskeletal stability remain unknown. Here, we created add1-deficient zebrafish using Tol2 transposon-mediated gene trapping and evaluated how add1 deficiency affected early hematopoiesis development. When add1 is lacking in zebrafish, both the primitive erythropoiesis and definitive hematopoiesis are compromised, and the primitive erythroblast cells are unable to develop into healthy erythrocytes. More significantly, the RNA sequencing results demonstrated that the p53 pathway is activated in the add1-depletion erythroblast cells, causing the erythroblasts to undergo apoptosis at the 14-somites stage and 24 hpf. Additionally, the anemic phenotype and apoptosis in add1-deficient embryos can be partially rescued by p53 insufficiency. Taken together, our findings show that add1 is critical for zebrafish erythropoiesis partially through the p53-mediated apoptotic pathway, which expands the regulatory role of Add1 for nuclear function. To better understand the nuclear-specific function of add1, we used RNA-seq to analyze transcriptome changes in gata1-EGFP positive cells sorted from add1-deficient embryos and siblings at 22 hpf, when the differences in the number of erythrocytes were not yet evident in add1 mutant embryos. Comparative gene expression profiling analysis of RNA-seq data for erythroblast of add1 mutant and siblings.