Hyperactivation of mTORC1 disrupts cellular homeostasis in cerebellar Purkinje cells

Yusuke Sakai,Atsu Aiba,Hiroyuki Sakagami,Tatsuya Maeda,Kazuki Nakao,Masahiro Fukaya,Kouichi Hashimoto,Hidetoshi Kassai,Hisako Nakayama,Masanobu Kano

doi:10.1038/s41598-019-38730-4

Abstract

Mammalian target of rapamycin (mTOR) is a central regulator of cellular metabolism. The importance of mTORC1 signaling in neuronal development and functions has been highlighted by its strong relationship with many neurological and neuropsychiatric diseases. Previous studies demonstrated that hyperactivation of mTORC1 in forebrain recapitulates tuberous sclerosis and neurodegeneration. In the mouse cerebellum, Purkinje cell-specific knockout of Tsc1/2 has been implicated in autistic-like behaviors. However, since TSC1/2 activity does not always correlate with clinical manifestations as evident in some cases of tuberous sclerosis, the intriguing possibility is raised that phenotypes observed in Tsc1/2 knockout mice cannot be attributable solely to mTORC1 hyperactivation. Here we generated transgenic mice in which mTORC1 signaling is directly hyperactivated in Purkinje cells. The transgenic mice exhibited impaired synapse elimination of climbing fibers and motor discoordination without affecting social behaviors. Furthermore, mTORC1 hyperactivation induced prominent apoptosis of Purkinje cells, accompanied with dysregulated cellular homeostasis including cell enlargement, increased mitochondrial respiratory activity, and activation of pseudohypoxic response. These findings suggest the different contributions between hyperactivated mTORC1 and Tsc1/2 knockout in social behaviors, and reveal the perturbations of cellular homeostasis by hyperactivated mTORC1 as possible underlying mechanisms of neuronal dysfunctions and death in tuberous sclerosis and neurodegenerative diseases.

Highlights

Mammalian target of rapamycin is an evolutionarily conserved protein kinase that acts as two functionally distinct complexes, termed mTORC1 and mTORC21. mTORC1 signaling serves as a central hub for the regulation of cellular metabolism, integrating various environmental stimuli such as growth hormones and amino acids[2]
We have shown that cortical microcephaly by hyperactivation of mTORC1 in the embryonic cortex is due to excessive apoptosis of cortical progenitors, accompanied by elevated expression of both hypoxia-inducible factor (HIF) −1α and its downstream gene[5]
Several animal models for mTOR hyperactivation in Purkinje cells have been generated by deleting Tsc[1] or Tsc[2] gene, a negative regulator of mTORC119,20

Summary

Introduction

Mammalian (or mechanistic) target of rapamycin (mTOR) is an evolutionarily conserved protein kinase that acts as two functionally distinct complexes, termed mTORC1 and mTORC21. mTORC1 signaling serves as a central hub for the regulation of cellular metabolism, integrating various environmental stimuli such as growth hormones and amino acids[2]. The Purkinje cell is innervated by multiple CFs and surplus CFs are gradually eliminated to establish mono-innervation in adulthood[10] Both motor coordination and synapse elimination are hallmarks of Purkinje cell functions, and many synaptic proteins are involved in these processes[10]. Purkinje cell-specific Tsc1/2 knockout mice exhibit abnormal behaviors in social interaction test, suggesting that aberrant activation of mTORC1 in Purkinje cells may be responsible for the onset of ASD-like symptoms. Human patients with N525S in TSC2 display severe symptoms of tuberous sclerosis without affecting TSC1/2 complex formation or GAP activity toward Rheb, whereas G1556S mutation impairs GAP activity with mild symptoms[13,14]. Our findings provide evidence that mTORC1 signaling in Purkinje cells is important for maintenance of cellular homeostasis

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Results

Conclusion