The conditional KO approach: Cre/Lox technology in human neurons.

ABSTRACT The use of human pluripotent stem cells to model human diseases has become a new standard in biomedical sciences. To this end, patient-derived somatic cells are studied in vitro to mimic human pathological conditions. Here, we describe an alternative experimental strategy, the ‘conditional KO approach’, which allows engineering disease-relevant mutations in pluripotent stem cells from healthy donors. In combination with the Cre/Lox technology, this strategy enables us to study the molecular causes of human diseases independent of the genetic background or of genetic alterations induced by clonal selection. As a proof-of-principle, we generated pluripotent stem cells with conditional loss-of-function mutations in the human STXBP1 gene that encodes Munc18-1. Using neurons derived from these cells, we show that heterozygous disruption of STXBP1 produces a specific and selective impairment in synaptic transmission that may account for the severe neurological disease caused by such mutations in human patients.