NudC L279P Mutation Destabilizes Filamin A by Inhibiting the Hsp90 Chaperoning Pathway and Suppresses Cell Migration.

Min Liu,Yiqing Lu,Tianhua Zhou,Jiaxing Feng,Chunxia Yang,Wei Liu,Yuehong Yang,Mingyang Yang,Wenwen Chen,Cheng Zhang,Xiaoyang Xu,Zhangqi Xu,Xiaoxia Sun,Wen Zhang

doi:10.3389/fcell.2021.671233

Abstract

Filamin A, the first discovered non-muscle actin filament cross-linking protein, plays a crucial role in regulating cell migration that participates in diverse cellular and developmental processes. However, the regulatory mechanism of filamin A stability remains unclear. Here, we find that nuclear distribution gene C (NudC), a cochaperone of heat shock protein 90 (Hsp90), is required to stabilize filamin A in mammalian cells. Immunoprecipitation-mass spectrometry and western blotting analyses reveal that NudC interacts with filamin A. Overexpression of human NudC-L279P (an evolutionarily conserved mutation in NudC that impairs its chaperone activity) not only decreases the protein level of filamin A but also results in actin disorganization and the suppression of cell migration. Ectopic expression of filamin A is able to reverse these defects induced by the overexpression of NudC-L279P. Furthermore, Hsp90 forms a complex with filamin A. The inhibition of Hsp90 ATPase activity by either geldanamycin or radicicol decreases the protein stability of filamin A. In addition, ectopic expression of Hsp90 efficiently restores NudC-L279P overexpression-induced protein stability and functional defects of filamin A. Taken together, these data suggest NudC L279P mutation destabilizes filamin A by inhibiting the Hsp90 chaperoning pathway and suppresses cell migration.

Highlights

Cell migration, a highly integrated multistep process, plays a critical role in diverse cellular and developmental processes, including the inflammatory response, tissue repair, embryogenesis, and cancer metastasis (Ridley et al, 2003; George et al, 2013)
To identify the candidate proteins participating in NudCmediated cell migration, we performed immunoprecipitation (IP) analysis with endogenous nuclear distribution gene C (NudC) followed by mass spectrometry (MS) in HeLa cells and found that approximately three hundred proteins appeared to associate with NudC (Supplementary Table 1, ranked based on relative abundance)
The results showed that F7 and F8 of filamin A appeared to interact with His-NudC (Figures 1G,H), suggesting that there is a direct interaction between NudC with both the 20-23 repeats (2132-2516 aa) and with repeat 24 (2517-2648 aa) of filamin A

Summary

Introduction

A highly integrated multistep process, plays a critical role in diverse cellular and developmental processes, including the inflammatory response, tissue repair, embryogenesis, and cancer metastasis (Ridley et al, 2003; George et al, 2013). Accumulating evidence indicates that the dynamic actin cytoskeleton spatially and temporally regulates cell migration. There exists a dynamically remodeling of branched actin filament network in lamellipodia, which is tightly regulated by several actin regulators, including actin-related protein 2/3 (Arp2/3), cofilin, profilin, and filamins (Blanchoin et al, 2000; Cameron et al, 2000; Skau and Waterman, 2015). Filamin A is widely expressed and plays an important role in cell migration regulation (Stossel et al, 2001; Feng and Walsh, 2004; Razinia et al, 2012; Kircher et al, 2015; Bandaru et al, 2019). A series of studies indicate that Asb (ankyrin repeat-containing protein with a suppressor of cytokine signaling box 2), a specific subunit of CRL5 (Cullin 5-RING E3 ubiquitin ligases) is involved in proteasomal degradation of filamin A (Heuze et al, 2008; Razinia et al, 2011, 2013; Lamsoul et al, 2013; Spinner et al, 2015), the regulation of filamin A stability remains poorly understood

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Conclusion