FishCOLler: generation of in vivo models of COL4A1/A2 small vessel disease in tractable zebrafish embryos to unravel patho-mechanisms and targetable molecules

Collagen IV a1/a2 is the major component of basement membranes, supporting key cellular processes, i.e. brain blood vessels stability. Not surprisingly, mutations affecting the genes (COL4A1,A2) cause a heterogenous disease that can affect multiple organs with varying severity and onset. Spontaneous or trauma-induced intracerebral haemorrhages (ICH) are common, causing brain cysts and epilepsy when occur prenatally. Cataract, retinal pathology, nephropathy and motor symptoms can also occur. Mutations can be inherited and affect only some family members or occur “de novo” in affected children. We currently lack complete understanding of the full disease spectrum and of the pathological significance of “candidate” COL4A variants continuously identified in undiagnosed patients. Fast and reliable research workflows for speedy variants’ validation in living organisms and large-scale mechanisms testing in patient-specific manner are necessary to improve differential diagnosis, provide clear guidelines preventing fatalities and develop therapies. While studies in mice were pivotal for disease identification, their lengthily, costly and often invasive protocols are inadequate to this aim. Here, I propose to generate disease models in the small freshwater fish, zebrafish, genetically similar to humans and suitable for rapid gene-editing and analysis in entire embryos. In see-through fish mutants we will assess brain, vessels, eyes and muscles pathology as well as ICH and epilepsy susceptibility via high-resolution microscopy and ad hoc assays we developed. Analysis of molecular alterations will contribute to unravel disease fingerprints and prospective targets. The project will generate a unique resource for rapid patient mutations testing in entire organisms towards speeding up therapy development.