Increased dopamine and norepinephrine levels with no difference in DA turnover in KO mice led us to hypothesize that FoxO1 is directly involved in CA synthesis in DA neurons. We found a significant increase in mRNA and protein levels of TH in DA neurons of KO mice, which was not detected in other brain regions (Fig. 7a–c and Supplementary Fig. 6a–c). Moreover, the phosphorylated form of TH was significantly increased in DA neuron regions of KO mice further suggesting the increase in TH activity and CA synthesis in KO mice (Fig. 7a–c and Supplementary Fig. 6a–c). Since the highly elevated TH expression was accompanied by an increase in dopamine and norepinephrine levels in KO mice, we speculated that FoxO1 might transcriptionally regulate TH expression. Indeed, sequence analysis revealed three potential FoxO1-binding sites including one conserved FoxO1-binding motif at the proximal region and two insulin-responsive elements (IREs) at the middle and distal regions within −3 kb of TH promoter (Fig. 7d)39. We therefore performed chromatin immonoprecipitaion (ChIP) assays to determine whether FoxO1 binds directly to these potential binding sites. Both in
