Human CLOCK enhances cognitive flexibility by altering cortical excitatory neuron function

A BAC containing human CLOCK gene with flanking regulatory region was transferred to mice. The humazied (HU) mice outperformed wildtype (WT) in a reversal learning task, and hence suggested enhanced cognitive flexibility in HU mice. IHC of frontal cortex suggested that the enhanced cognitive flexibility in HU mice might result from the increased neuronal density, dendritic arborization, and spine density. In order to determine the molecular mechanism, we conducted single-nuclei RNA-seq on frontal cortex from postnatal 7 day (P7) and P56 mice. In the excitatory neuron, genes that upregulated in HU mice were enriched for dendritic growth (e.g. Tenm2 and Flrt2), spine formation (e.g. Cntn5 and Sorcs2), and energy metabolism (e.g. Suclg1 and Cox8a). P7 and P56 of humanized (CLOCKhum;Clock-/-, HU), knockout (Clock-/-, KO), and wildtype (Clock+/+, WT) mice were harvested rostral part of frontal cortex which is corresponding to the cortex part of Coronal level 22-36 in Allen Brain Atlas: Reference Atlas, Version 2 (2011), and extracted single nuclei for sequencing.