Activation of mitochondrial TUFM ameliorates metabolic dysregulation through coordinating autophagy induction

Dasol Kim,Jin Young Kim,Ho Jeong Kwon,Hui-Yun Hwang,Jong Shin Yoo,Eun Sun Ji

doi:10.1038/s42003-020-01566-0

Abstract

Disorders of autophagy, a key regulator of cellular homeostasis, cause a number of human diseases. Due to the role of autophagy in metabolic dysregulation, there is a need to identify autophagy regulators as therapeutic targets. To address this need, we conducted an autophagy phenotype-based screen and identified the natural compound kaempferide (Kaem) as an autophagy enhancer. Kaem promoted autophagy through translocation of transcription factor EB (TFEB) without MTOR perturbation, suggesting it is safe for administration. Moreover, Kaem accelerated lipid droplet degradation in a lysosomal activity-dependent manner in vitro and ameliorated metabolic dysregulation in a diet-induced obesity mouse model. To elucidate the mechanism underlying Kaem’s biological activity, the target protein was identified via combined drug affinity responsive target stability and LC–MS/MS analyses. Kaem directly interacted with the mitochondrial elongation factor TUFM, and TUFM absence reversed Kaem-induced autophagy and lipid degradation. Kaem also induced mitochondrial reactive oxygen species (mtROS) to sequentially promote lysosomal Ca2+ efflux, TFEB translocation and autophagy induction, suggesting a role of TUFM in mtROS regulation. Collectively, these results demonstrate that Kaem is a potential therapeutic candidate/chemical tool for treating metabolic dysregulation and reveal a role for TUFM in autophagy for metabolic regulation with lipid overload.

Highlights

Disorders of autophagy, a key regulator of cellular homeostasis, cause a number of human diseases
The challenges associated with preparing modified chemicals engendered the development of alternative methods in which unmodified compounds are used such as drug affinity response target stability (DARTS), which exploits changes in the protease susceptibility of target proteins upon chemical binding[19,20,21]
acridine orange (AO) staining is a well-known assay to examine the function and integrity of lysosome, which is used to evaluate the status of autophagic flux[27,28] Correlation of the readout with AO fluorescence was validated via measuring the intensity of cells that are stained with different concentration of the staining (Supplementary Fig. 1a), and via checking that a positive control indatraline, which enhances lysosomal acidity[29], increased AO intensity 1.2-fold, and a negative control bafilomycin A1, which inhibits acidic lysosome by perturbing proton channel[24], decreased AO intensity 0.7-fold (Supplementary Fig. 1b–d)

Summary

Introduction

A key regulator of cellular homeostasis, cause a number of human diseases. Kaem induced mitochondrial reactive oxygen species (mtROS) to sequentially promote lysosomal Ca2+ efflux, TFEB translocation and autophagy induction, suggesting a role of TUFM in mtROS regulation These results demonstrate that Kaem is a potential therapeutic candidate/chemical tool for treating metabolic dysregulation and reveal a role for TUFM in autophagy for metabolic regulation with lipid overload. We report that Kaem has a role in regulating metabolic fitness by enhancing autophagy This effect does not involve perturbation of the major autophagy regulatory factor, MTOR, which has a crucial role in cellular growth, suggesting that MTOR-related side effects can be avoided. We leveraged DARTS analysis with an LC–MS/MS quantitative proteomics approach as a label-free method to identify target proteins of the autophagy-enhancing natural compound Kaem Using this approach, we identified the mitochondrial translation factor TUFM as a target of Kaem, and the physical interaction was confirmed in vitro. LC–MS/MS analysis upon Kaem treatment provides insight on TUFM in mitochondrial reactive oxygen species (mtROS) regulation and a mechanism underlying Kaem-induced autophagy

Methods

Results

Conclusion