GlialbetaPixis essential for blood vessel integrity in the zebrafish brain

The formation of blood-brain barrier and vascular integrity depends on the coordinative development of different cell types in the brain. Previous studies have shown that zebrafishbubblehead(bbh) mutant, which has mutation in thebetaPixlocus, develops spontaneous intracerebral hemorrhage during early development. However, it remains unclear in which brain cellsbetaPixmay function. Here, we established a highly efficient conditional knockout method in zebrafish by using homology-directed repair (HDR)-mediated knockin and knockout technology, and generatedbetaPixconditional trap (betaPix^ct) allele in zebrafish. We found thatbetaPixin glia, but neither neurons, endothelial cells, nor pericytes, was critical for glial and vascular development and integrity, thus contributing to the formation of blood-brain barrier. Single-cell transcriptome profiling revealed that microtubule aggregation signaling stathmins and pro-angiogenic transcription factorsZfhx3/4were down-regulated in glial and neuronal progenitors, and further genetic analysis suggested that betaPix acted upstream on the PAK1-Stathmin and Zfhx3/4-Vegfaa signaling to regulate glia migration and vascular integrity. Therefore, this work reveals that glialbetaPixplays an important role in brain vascular integrity in zebrafish embryos and possibly human cells.