Bulk RNA-sequencing of pooled 53 hpf wild-type and iqgap1-/- hemorrhage-positive mutant embryos

ABSTRACT: Defects in multiple cell-signaling molecules lead to defects of vascular integrity given the need for fine-tuned regulation of the cell adhesion complexes. Signaling adaptor proteins play a crucial in cell signaling by promoting interactions between effector proteins and even enabling integration of different pathways. IQGAP1 is a conserved signaling adaptor known for its roles in cell adhesion, cancer and for other cell biological effects. We engineered a zebrafish null mutant in iqgap1 gene by introducing an 11-bp deletion by CRISPR/Cas9 and characterized its phenotype. Homozygous mutants exhibit severe brain hemorrhage and morphological abnormalities, which are ultimately lethal in about 30-40% of cases, whereas the other embryos survive to adulthood. We assessed retinal vasculature in 12 to 16-month-old adults and, contrary to the human patient observations, did not find any find any differences between wild-type and iqgap1-/- mutant zebrafish. Expression pattern of iqgap1 strongly overlapped with the vascular marker fli1a expression but more broadly present in tissues such as muscle, branchial arches, caudal hematopoietic tissue. Re-analysis of single-cell RNA-seq data extended the known expression pattern of iqgap1 to include mesenchyme, periderm, neural cell types, granulocytes and macrophages. Strikingly, iqgap1 showed a striking co-localization with fli1a in the brain blood vessels, whose disruption likely causes the brain hemorrhage. RNA sequencing-based comparison of whole hemorrhage-positive iqgap1-/- embryo pools with wild-type embryos at 54 hpf shortly after the onset of hemorrhage identified 1283 differentially regulated genes. The most striking feature of this dataset was up-regulation of hematopoietic markers especially those of erythrocytes, neutrophils, mast cells and HSPCs (hematopoietic stem and progenitor cells) but macrophages were down-regulated. We have confirmed that erythrocytes up-regulation is linked to hemorrhage. By contrast, mast cells, neutrophils are up-regulated and macrophages are reduced in iqgap1-/- embryos with or without hemorrhage.

ORGANISM(S): Danio rerio

PROVIDER: GSE330159 | GEO | 2026/05/12

REPOSITORIES: GEO