Emma Andersson

Title: In utero nano-injection as a tool to manipulate murine gene expression

Abstract: Major advances in genomics and transcriptomics have rapidly advanced our knowledge into genes and mechanisms that may be controlling aspects of health and disease. However, in vivo validation has lagged behind. To address this, we adapted in utero nano-injection to target and manipulate gene expression in embryonic tissues during or after gastrulation. One iteration of the technique, which we dubbed NEPTUNE (neural plate targeting by in utero nano-injection) rapidly and flexibly transduces the neural plate with virus prior to neurulation, and can thus manipulate the future nervous system. Stable integration in >95% of cells in the brain enabled long-term over-expression, and conditional expression can be achieved using cell-type specific promoters. In the absence of conditional expression, this method broadly targets ectodermal derivatives including epidermis and neural crest-derived mesenchyme, as well as ectodermal placode-derive organs such as the eye and inner ear. The method can recapitulate established mouse knockouts. Finally, we used NEPTUNE to investigate the function of Sptn2, mutations in which are associated with Spinocerebellar ataxia type 5. Sptbn2 knockout mice exhibit motor deficits, similar to patients. However, Sptbn2 knockdown with NEPTUNE induced dose-dependent embryonic lethal defects in neural tube, embryonic turning, and abdominal wall closure, previously unreported functions for Sptbn2. In utero nano-injection thus offers a rapid and cost-effective technique to test gene function in developing embryos and can reveal phenotypes incompatible with life.