Glucocorticoids rapidly inhibit cell migration through a novel, non-transcriptional HDAC6 pathway.

Stephen Kershaw,Egor Zindy,Magnus Rattray,Mudassar Iqbal,Laura C Matthews,Andrew Brass,David G Spiller,Gareth Kitchen,David W Ray,David J Morgan,Claus Jorgensen,Tracy Hussell,Chris M Sanderson,James Boyd

doi:10.1242/jcs.242842

Abstract

ABSTRACTGlucocorticoids (GCs) act through the glucocorticoid receptor (GR, also known as NR3C1) to regulate immunity, energy metabolism and tissue repair. Upon ligand binding, activated GR mediates cellular effects by regulating gene expression, but some GR effects can occur rapidly without new transcription. Here, we show that GCs rapidly inhibit cell migration, in response to both GR agonist and antagonist ligand binding. The inhibitory effect on migration is prevented by GR knockdown with siRNA, confirming GR specificity, but not by actinomycin D treatment, suggesting a non-transcriptional mechanism. We identified a rapid onset increase in microtubule polymerisation following GC treatment, identifying cytoskeletal stabilisation as the likely mechanism of action. HDAC6 overexpression, but not knockdown of αTAT1, rescued the GC effect, implicating HDAC6 as the GR effector. Consistent with this hypothesis, ligand-dependent cytoplasmic interaction between GR and HDAC6 was demonstrated by quantitative imaging. Taken together, we propose that activated GR inhibits HDAC6 function, and thereby increases the stability of the microtubule network to reduce cell motility. We therefore report a novel, non-transcriptional mechanism whereby GCs impair cell motility through inhibition of HDAC6 and rapid reorganization of the cell architecture.This article has an associated First Person interview with the first author of the paper.

Highlights

Glucocorticoids (GCs) are steroid hormones that regulate a range of biological functions essential for life, including normal homeostasis, glucose metabolism, resolution of inflammation and development (McMaster et al, 2008; Tu et al, 2018; Tanaka et al, 2017)
Deacetylation of α-tubulin is catalysed by histone deacetylase-6 (HDAC6) and modulation of HDAC6 activity impacts cell migration by altering the dynamics of the microtubule network (Hubbert et al, 2002; Zhang et al, 2007; Shi et al, 2015)
glucocorticoid receptor (GR) agonists and antagonists inhibit cell migration Dexamethasone, a synthetic GR agonist, potently inhibits the migration of A549 cells tracked by immunofluorescence imaging of GFP-labelled histone 2B (H2B) (Fig. 1A), causing a marked reduction in total displacement and step length compared to vehicle-treated controls (Fig. 1B)

Summary

Introduction

Glucocorticoids (GCs) are steroid hormones that regulate a range of biological functions essential for life, including normal homeostasis, glucose metabolism, resolution of inflammation and development (McMaster et al, 2008; Tu et al, 2018; Tanaka et al, 2017). GCs exert their biological effects through the ubiquitously expressed glucocorticoid receptor (GR; known as NR3C1), a ligand-inducible transcription factor of the nuclear hormone receptor superfamily (Hollenberg et al, 1985). GCs are known to inhibit the migration of various cell types, yet with an unrecognised mechanism of action (Fietz et al, 2017; Murakami et al, 1998). Deacetylation of α-tubulin is catalysed by histone deacetylase-6 (HDAC6) and modulation of HDAC6 activity impacts cell migration by altering the dynamics of the microtubule network (Hubbert et al, 2002; Zhang et al, 2007; Shi et al, 2015). Overexpression of HDAC6 increases cell motility by regulating microtubule-dependent migration (Ridley et al, 2003; Wu and Bezanilla, 2018). HDAC6 deacetylates heat-shock protein-90 (Hsp90), which is vital for GR maturation and maintaining the receptor in a ligand-binding state

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