AbstractThe KH-type splicing regulatory protein (KHSRP) is an RNA-binding protein linked to decay of mRNAs with AU-rich elements. KHSRP was previously shown to destabilize Gap43 mRNA and decrease neurite growth in cultured embryonic neurons. Here, we have tested functions of KHSRP in vivo. We find upregulation of 1460 mRNAs in neocortex of adult Khsrp^−/− mice, of which 527 bind to KHSRP with high specificity. These KHSRP targets are involved in pathways for neuronal morphology, axon guidance, neurotransmission and long-term memory. Khsrp^−/− mice show increased axon growth and dendritic spine density in vivo. Neuronal cultures from Khsrp^−/− mice show increased axon and dendrite growth and elevated KHSRP-target mRNAs, including subcellularly localized mRNAs. Furthermore, neuron-specific knockout of Khsrp confirms these are from neuron-intrinsic roles of KHSRP. Consistent with this, neurons in the hippocampus and infralimbic cortex of Khsrp^−/− mice show elevations in frequency of miniature excitatory postsynaptic currents. The Khsrp^−/− mice have deficits in trace conditioning and attention set-shifting tasks compared Khsrp^+/+ mice, indicating impaired prefrontal- and hippocampal-dependent memory consolidation with loss of KHSRP. Overall, these results indicate that deletion of KHSRP impairs neuronal development resulting in alterations in neuronal morphology and function by changing post-transcriptional control of neuronal gene expression.