Phosphoproteomic of the acetylcholine pathway enables discovery of the PKC-β-PIX-Rac1-PAK cascade as a stimulatory signal for aversive learning

Yukie Yamahashi, You Hsin Lin, Akihiro Mouri, Sho Iwanaga, Kazuhiro Kawashima, Yuya Tokumoto, Yo Watanabe, Md Omar Faruk, Xinjian Zhang, Daisuke Tsuboi, Takashi Nakano, Naoaki Saito, Taku Nagai, Kiyofumi Yamada, Kozo Kaibuchi

Research output: Contribution to journalArticlepeer-review

Abstract

Acetylcholine is a neuromodulator critical for learning and memory. The cholinesterase inhibitor donepezil increases brain acetylcholine levels and improves Alzheimer’s disease (AD)-associated learning disabilities. Acetylcholine activates striatal/nucleus accumbens dopamine receptor D2-expressing medium spiny neurons (D2R-MSNs), which regulate aversive learning through muscarinic receptor M1 (M1R). However, how acetylcholine stimulates learning beyond M1Rs remains unresolved. Here, we found that acetylcholine stimulated protein kinase C (PKC) in mouse striatal/nucleus accumbens. Our original kinase-oriented phosphoproteomic analysis revealed 116 PKC substrate candidates, including Rac1 activator β-PIX. Acetylcholine induced β-PIX phosphorylation and activation, thereby stimulating Rac1 effector p21-activated kinase (PAK). Aversive stimulus activated the M1R-PKC-PAK pathway in mouse D2R-MSNs. D2R-MSN-specific expression of PAK mutants by the Cre-Flex system regulated dendritic spine structural plasticity and aversive learning. Donepezil induced PAK activation in both accumbal D2R-MSNs and in the CA1 region of the hippocampus and enhanced D2R-MSN-mediated aversive learning. These findings demonstrate that acetylcholine stimulates M1R-PKC-β-PIX-Rac1-PAK signaling in D2R-MSNs for aversive learning and imply the cascade’s therapeutic potential for AD as aversive learning is used to preliminarily screen AD drugs.

Original languageEnglish
JournalMolecular Psychiatry
DOIs
Publication statusAccepted/In press - 2022

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Cellular and Molecular Neuroscience
  • Psychiatry and Mental health

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