A TFEB nuclear export signal integrates amino acid supply and glucose availability

Linxin Li,Colin R Goding,Katherine L B Borden,Georgina Berridge,Panagis Filippakopoulos,Roman Fischer,Laurent Volpon,Kaochin Ngeow,Sarah Andrews,Hans J Friedrichsen,Sarah Picaud

doi:10.1038/s41467-018-04849-7

Abstract

How cells coordinate the response to fluctuating carbon and nitrogen availability required to maintain effective homeostasis is a key issue. Amino acid limitation that inactivates mTORC1 promotes de-phosphorylation and nuclear translocation of Transcription Factor EB (TFEB), a key transcriptional regulator of lysosome biogenesis and autophagy that is deregulated in cancer and neurodegeneration. Beyond its cytoplasmic sequestration, how TFEB phosphorylation regulates its nuclear-cytoplasmic shuttling, and whether TFEB can coordinate amino acid supply with glucose availability is poorly understood. Here we show that TFEB phosphorylation on S142 primes for GSK3β phosphorylation on S138, and that phosphorylation of both sites but not either alone activates a previously unrecognized nuclear export signal (NES). Importantly, GSK3β is inactivated by AKT in response to mTORC2 signaling triggered by glucose limitation. Remarkably therefore, the TFEB NES integrates carbon (glucose) and nitrogen (amino acid) availability by controlling TFEB flux through a nuclear import-export cycle.

Highlights

How cells coordinate the response to fluctuating carbon and nitrogen availability required to maintain effective homeostasis is a key issue
As most studies examine the steady state location of Transcription Factor EB (TFEB), we established a stably expressed GFP-reporter system in which the dynamics of TFEB cytoplasmic-nuclear shuttling could be examined in realtime by using MCF7 cells in which TFEB-GFP was under the control of a doxycycline-inducible promoter
Examination of TFEB-GFP under these conditions revealed that TFEB subcellular localization was highly dynamic; over the course of 20 min TFEB in some cells was seen to accumulate in the nucleus and return to the cytoplasm (Fig. 1b; Supplementary Movie 1), presumably indicating that TFEB responds to changing intracellular nutrient availability even within cells grown in a nutrient rich environment

Summary

Introduction

How cells coordinate the response to fluctuating carbon and nitrogen availability required to maintain effective homeostasis is a key issue. In addition to promoting lysosome biogenesis in response to amino acid limitation, TFEB can enhance the integrated stress response mediated by ATF416 and acts as a nexus for nutrient sensing and resolution of any supply-demand disequilibrium. It is a key effector of the beneficial effects of exercise by controlling metabolic flexibility in muscle[17], protects against inflammation-mediated atherosclerosis[18], and neurodegenerative disease[13,19–21] and is deregulated in cancer[22]. We found that TFEB has a regulated nuclear export signal (NES) in which phosphorylation at the ERK/mTORC1 phosphorylation site at S142 primed for phosphorylation by GSK3β at S138. The results establish that nuclear export is a critical nexus for regulation of TFEB subcellular localization

Methods

Results

Conclusion