Phosphoproteomic of the Acetylcholine Pathway Enables Discovery of the PKC-β-PIX-Rac1-PAK Cascade As a Stimulatory Signal for Aversive Learning

Overview

Journal Mol Psychiatry

Specialties Molecular Biology
Psychiatry

Date 2022 Jun 6

PMID 35665767

Authors

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

Affiliations

Soon will be listed here.

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.

Citing Articles

Role of adenosine in the pathophysiology and treatment of attention deficit hyperactivity disorder.

Jia Q, Tan H, Li T, Duan X Purinergic Signal. 2024; .

PMID: 39480600 DOI: 10.1007/s11302-024-10059-2.

GSK3β Substrate-competitive Inhibitors Regulate the gut Homeostasis and Barrier Function to Inhibit Neuroinflammation in Scopolamine-induced Alzheimer's Disease Model Mice.

Zhang L, Jiang Z, Hu S, Ni H, Zhao Y, Tan X Inflammation. 2024; .

PMID: 39180577 DOI: 10.1007/s10753-024-02133-z.

Multiple cholinergic receptor subtypes coordinate dual modulation of acetylcholine on anterior and posterior paraventricular thalamic neurons.

Ye Q, Nunez J, Zhang X J Neurochem. 2024; 168(6):995-1018.

PMID: 38664195 PMC: 11136594. DOI: 10.1111/jnc.16115.

KANPHOS: Kinase-associated neural phospho-signaling database for data-driven research.

Kannon T, Murashige S, Nishioka T, Amano M, Funahashi Y, Tsuboi D Front Mol Neurosci. 2024; 17:1379089.

PMID: 38628370 PMC: 11018961. DOI: 10.3389/fnmol.2024.1379089.

Neuromodulator regulation and emotions: insights from the crosstalk of cell signaling.

Tsuboi D, Nagai T, Yoshimoto J, Kaibuchi K Front Mol Neurosci. 2024; 17:1376762.

PMID: 38516040 PMC: 10954900. DOI: 10.3389/fnmol.2024.1376762.

References

Barker L, Glick S, Green J, Khandelwal J . Acetylcholine metabolism in the rat hippocampus and striatum following one-trial passive training. Neuropharmacology. 1982; 21(2):183-5. DOI: 10.1016/0028-3908(82)90160-5. View

Yan Z, Flores-Hernandez J, Surmeier D . Coordinated expression of muscarinic receptor messenger RNAs in striatal medium spiny neurons. Neuroscience. 2001; 103(4):1017-24. DOI: 10.1016/s0306-4522(01)00039-2. View

Cox J, Mann M . MaxQuant enables high peptide identification rates, individualized p.p.b.-range mass accuracies and proteome-wide protein quantification. Nat Biotechnol. 2008; 26(12):1367-72. DOI: 10.1038/nbt.1511. View

Newey S, Velamoor V, Govek E, Van Aelst L . Rho GTPases, dendritic structure, and mental retardation. J Neurobiol. 2005; 64(1):58-74. DOI: 10.1002/neu.20153. View

Funahashi Y, Ariza A, Emi R, Xu Y, Shan W, Suzuki K . Phosphorylation of Npas4 by MAPK Regulates Reward-Related Gene Expression and Behaviors. Cell Rep. 2019; 29(10):3235-3252.e9. DOI: 10.1016/j.celrep.2019.10.116. View

Dickstein D, Dickstein D, Janssen W, Hof P, Glaser J, Rodriguez A . Automatic Dendritic Spine Quantification from Confocal Data with Neurolucida 360. Curr Protoc Neurosci. 2016; 77:1.27.1-1.27.21. PMC: 5113738. DOI: 10.1002/cpns.16. View

Surti T, Huang L, Jan Y, Jan L, Cooper E . Identification by mass spectrometry and functional characterization of two phosphorylation sites of KCNQ2/KCNQ3 channels. Proc Natl Acad Sci U S A. 2005; 102(49):17828-33. PMC: 1297712. DOI: 10.1073/pnas.0509122102. View

Sugimoto H . Donepezil hydrochloride: a treatment drug for Alzheimer's disease. Chem Rec. 2002; 1(1):63-73. DOI: 10.1002/1528-0691(2001)1:1<63::AID-TCR9>3.0.CO;2-J. View

Gerfen C, Engber T, Mahan L, Susel Z, Chase T, Monsma Jr F . D1 and D2 dopamine receptor-regulated gene expression of striatonigral and striatopallidal neurons. Science. 1990; 250(4986):1429-32. DOI: 10.1126/science.2147780. View

10.

Dolan B, Duron S, Campbell D, Vollrath B, Rao B, Ko H . Rescue of fragile X syndrome phenotypes in Fmr1 KO mice by the small-molecule PAK inhibitor FRAX486. Proc Natl Acad Sci U S A. 2013; 110(14):5671-6. PMC: 3619302. DOI: 10.1073/pnas.1219383110. View

11.

Quirion R, Araujo D, Regenold W, Boksa P . Characterization and quantitative autoradiographic distribution of [3H]acetylcholine muscarinic receptors in mammalian brain. Apparent labelling of an M2-like receptor sub-type. Neuroscience. 1989; 29(2):271-89. DOI: 10.1016/0306-4522(89)90057-2. View

12.

QUASTEL J, Tennenbaum M, Wheatley A . Choline ester formation in, and choline esterase activities of, tissues in vitro. Biochem J. 1936; 30(9):1668-81. PMC: 1263240. DOI: 10.1042/bj0301668. View

13.

Zhao Z, Manser E . PAK and other Rho-associated kinases--effectors with surprisingly diverse mechanisms of regulation. Biochem J. 2004; 386(Pt 2):201-14. PMC: 1134783. DOI: 10.1042/BJ20041638. View

14.

Nishioka T, Amano M, Funahashi Y, Tsuboi D, Yamahashi Y, Kaibuchi K . In Vivo Identification of Protein Kinase Substrates by Kinase-Oriented Substrate Screening (KIOSS). Curr Protoc Chem Biol. 2019; 11(1):e60. DOI: 10.1002/cpch.60. View

15.

Zhao Z, Manser E, Chen X, Chong C, Leung T, Lim L . A conserved negative regulatory region in alphaPAK: inhibition of PAK kinases reveals their morphological roles downstream of Cdc42 and Rac1. Mol Cell Biol. 1998; 18(4):2153-63. PMC: 121452. DOI: 10.1128/MCB.18.4.2153. View

16.

Hoebel B, Avena N, Rada P . Accumbens dopamine-acetylcholine balance in approach and avoidance. Curr Opin Pharmacol. 2007; 7(6):617-27. PMC: 2727589. DOI: 10.1016/j.coph.2007.10.014. View

17.

Kutys M, Yamada K . An extracellular-matrix-specific GEF-GAP interaction regulates Rho GTPase crosstalk for 3D collagen migration. Nat Cell Biol. 2014; 16(9):909-17. PMC: 4150836. DOI: 10.1038/ncb3026. View

18.

Mouri A, Sasaki A, Watanabe K, Sogawa C, Kitayama S, Mamiya T . MAGE-D1 regulates expression of depression-like behavior through serotonin transporter ubiquitylation. J Neurosci. 2012; 32(13):4562-80. PMC: 6622051. DOI: 10.1523/JNEUROSCI.6458-11.2012. View

19.

Faruk M, Tsuboi D, Yamahashi Y, Funahashi Y, Lin Y, Ahammad R . Muscarinic signaling regulates voltage-gated potassium channel KCNQ2 phosphorylation in the nucleus accumbens via protein kinase C for aversive learning. J Neurochem. 2021; 160(3):325-341. DOI: 10.1111/jnc.15555. View

20.

Nakajo K, Kubo Y . Protein kinase C shifts the voltage dependence of KCNQ/M channels expressed in Xenopus oocytes. J Physiol. 2005; 569(Pt 1):59-74. PMC: 1464213. DOI: 10.1113/jphysiol.2005.094995. View