Smad5 acts as an intracellular pH messenger and maintains bioenergetic homeostasis

Yujiang Fang,Yi Yang,Yi Hui,Bin Xiang,Daniel Terheyden-Keighley,Mengtao Xiao,Bowen Zhang,Yanhua Du,Lin Ma,Hexi Feng,Quanbin Zhang,Zhongliang Liu,Xin Zhang,Guohua Jin,Yanyan Yu,Yuanyuan Zhang,Yuzheng Zhao,Lei Shi,Xiaobai Zhang,Cizhong Jiang,Zhenyu Chen,Beibei Fan,Xiangjie Xu,Ling Liu,Yiran Wang,Yang Liu,Xiaoqing Zhang

doi:10.1038/cr.2017.85

Abstract

Both environmental cues and intracellular bioenergetic states profoundly affect intracellular pH (pHi). How a cell responds to pHi changes to maintain bioenergetic homeostasis remains elusive. Here we show that Smad5, a well-characterized downstream component of bone morphogenetic protein (BMP) signaling responds to pHi changes. Cold, basic or hypertonic conditions increase pHi, which in turn dissociates protons from the charged amino acid clusters within the MH1 domain of Smad5, prompting its relocation from the nucleus to the cytoplasm. On the other hand, heat, acidic or hypotonic conditions decrease pHi, blocking the nuclear export of Smad5, and thus causing its nuclear accumulation. Active nucleocytoplasmic shuttling of Smad5 induced by environmental changes and pHi fluctuation is independent of BMP signaling, carboxyl terminus phosphorylation and Smad4. In addition, ablation of Smad5 causes chronic and irreversible dysregulation of cellular bioenergetic homeostasis and disrupted normal neural developmental processes as identified in a differentiation model of human pluripotent stem cells. Importantly, these metabolic and developmental deficits in Smad5-deficient cells could be rescued only by cytoplasmic Smad5. Cytoplasmic Smad5 physically interacts with hexokinase 1 and accelerates glycolysis. Together, our findings indicate that Smad5 acts as a pHi messenger and maintains the bioenergetic homeostasis of cells by regulating cytoplasmic metabolic machinery.

Highlights

Given that most biological reactions are pH depen-HCO3− anion exchangers which acidify the cells [2]
We unexpectedly found that GFP-Smad5 diffused from nucleus to cytoplasm within several minutes when the cells were placed at 25 °C (Figure 1A, Supplementary information, Figure S1A-S1D, Movie S1)
Smad5 is located both in the cytoplasm and nucleus; Smad1 is largely distributed in the nucleus; and Smad8 is mostly cytoplasmic

Summary

Introduction

Given that most biological reactions are pH depen-HCO3− anion exchangers which acidify the cells [2]. Upon BMP or TGFβ stimulation, cytosolic R-Smad members, including Smad, Smad, Smad, Smad and Smad, become phosphorylated by the type I receptors at the most C-terminal SSXS motif [6, 7] These phosphorylated R-Smads oligomerise with Smad and translocate into nucleus for gene transcriptional regulation [8]. Smad has been identified as playing a non-transcriptional role in regulating mitochondrial dynamics and function. This is facilitated by complexing with Mitofusin 2, Rab and Ras interactor 1 [12], suggesting a noncanonical role of the component within the BMP and TGFβ pathways in regulating metabolic processes. Smad deficiency results in abnormally swollen mitochondria and apoptosis in cardiomyocytes, indicating that it might be involved in energy metabolism [13]

Methods

Results

Conclusion