High-throughput sensitive screening of small molecule modulators of microexon alternative splicing using dual Nano and Firefly luciferase reporters

Disruption of alternative splicing frequently causes or contributes to human diseases and disorders. Consequently, there is a need for efficient and sensitive reporter assays capable of screening chemical libraries for compounds with efficacy in modulating important splicing events. Here, we describe a screening workflow employing dual Nano and Firefly luciferase alternative splicing reporters that affords highly efficient, sensitive, and linear detection of small molecule responses. Applying this system to a screen of ~95,000 small molecules, we identify compounds that selectively activate or repress a neuronal microexon network that is frequently disrupted in autism and overexpressed in neuroendocrine cancers. Remarkably, among the most potent and selective activating compounds are histone deacetylase (HDAC) inhibitors. We thus describe a high-throughput screening system for candidate splicing therapeutics, a resource of small molecule modulators of microexons, and insight into the mode of action and potential utility of HDAC inhibitors in the context of neurological disorders.