Deciphering the role of importin alpha in mammalian central neurons - Hippocampal gene expression profiling in importin alpha5-deleted mice

Introduction: The mechanisms governing synaptonuclear transport, and the transcriptional pathways that are at the core of neuronal disorders are poorly understood. The importin family of nuclear import factors has pivotal roles in such pathways, due to their involvement in intracellular transport from both synapse to soma and cytoplasm to nucleus. Mammals express six different isoforms of importin a, which bind an Importin ß to form transport complexes for specific cargoes. Methods: We addressed the roles of importin a isoforms in mammalian CNS by evaluation of a systematic series of importin a knockout mice. A comprehensive battery of behavioral tests was used to investigate spontaneous and novelty-induced locomotion, basal ganglia function, motor coordination, neuromuscular integration, anxiety-related behaviors and memory. Subsequent analyses using acute-brain slice electrophysiology, RNA-seq, bioinformatics, imaging and pharmacology was used in order to delineate the contribution of genes and signaling pathways involved in the identified phenotype and screen for new therapeutic approaches. Results and Discussion: During the course of comprehensive behavioral profiling of importin a mouse mutants, we identified an importin a knockout (KO) mouse with significantly reduced anxiety levels. Electrophysiological recordings in acute hippocampal slices showed a reduced short-term and presynaptic plasticity (paired-pulse facilitation deficit, lower PTP), while LTP was not affected in these mutants. We then performed transcriptional profiling (RNA-seq) from hippocampal extracts of wild type versus mutant animals after exposure to an anxiogenic drug. Computational analysis revealed differences in the expression of genes and the involvement of transcription factors. Drugs that activate LXR or the Sphingosine 1 phosphate receptor have robust anxiolytic effects, while a Sphk1 blocker reverses anxiolysis in the importin a5 knockout mouse. Thus, importin a5 influences anxiety by regulating nuclear import in neurons. Overall design: RNA-seq performed on: 4 experimental groups, 3 animals per each vehicle group and replicate, 3 replicates of the entire set - Exception in the FG7142-treated Importin a5-/- group (n=2) where 1 animal was identified outlier and discarded from the analysis