Interaction between Salt-inducible Kinase 2 (SIK2) and p97/Valosin-containing Protein (VCP) Regulates Endoplasmic Reticulum (ER)-associated Protein Degradation in Mammalian Cells

Fu-Chia Yang,Ya-Huei Lin,Wei-Hao Chen,Jing-Yi Huang,Hsin-Yun Chang,Su-Hui Su,Hsiao-Ting Wang,Chun-Yi Chiang,Pang-Hung Hsu,Ming-Daw Tsai,Bertrand Chin-Ming Tan,Sheng-Chung Lee

doi:10.1074/jbc.m113.492199

Abstract

Salt-inducible kinase 2 (SIK2) is an important regulator of cAMP response element-binding protein-mediated gene expression in various cell types and is the only AMP-activated protein kinase family member known to interact with the p97/valosin-containing protein (VCP) ATPase. Previously, we have demonstrated that SIK2 can regulate autophagy when proteasomal function is compromised. Here we report that physical and functional interactions between SIK2 and p97/VCP underlie the regulation of endoplasmic reticulum (ER)-associated protein degradation (ERAD). SIK2 co-localizes with p97/VCP in the ER membrane and stimulates its ATPase activity through direct phosphorylation. Although the expression of wild-type recombinant SIK2 accelerated the degradation and removal of ERAD substrates, the kinase-deficient variant conversely had no effect. Furthermore, down-regulation of endogenous SIK2 or mutation of the SIK2 target site on p97/VCP led to impaired degradation of ERAD substrates and disruption of ER homeostasis. Collectively, these findings highlight a mechanism by which the interplay between SIK2 and p97/VCP contributes to the regulation of ERAD in mammalian cells.

Highlights

Salt-inducible kinase 2 (SIK2) is the only AMPK family kinase that interacts with p97/valosin-containing protein (VCP) albeit without recognized functional consequence
Because ATPase activity of p97/VCP is essential for the retrotranslocation of misfolded proteins from endoplasmic reticulum (ER) to cytosol during ER-associated protein degradation (ERAD), the extent of which can be assessed by the residual levels of CD3␦ in the microsomal fraction, we examined the roles of SIK2 in this functional aspect
We demonstrated that SIK2 serves as an additional layer of ERAD regulation through its interaction with p97/VCP

Summary

Background

SIK2 is the only AMPK family kinase that interacts with p97/VCP albeit without recognized functional consequence. During ERAD, p97/VCP forms a complex with Ufd1-Npl heterodimer that plays a key role in extracting proteins from ER to the cytosolic proteasomal degradation machinery [23, 28] This process represents a critical determinant for degrading misfolded proteins and unassembled polypeptides of protein complexes in the ER such as the T cell antigen receptor CD3␦ subunit [29] as well as regulating the homeostasis of normal ER-resident proteins (30 –33). SIK2 interacts with p97/VCP and confers activation of its ATPase activity and ERAD function through mediating Ser-770 phosphorylation. Taken together, these results extended the known cellular roles of SIK2 to a critical function in ERAD

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION