Endothelial nuclear lamina is not required for glucocorticoid receptor nuclear import but does affect receptor-mediated transcription activation

Abstract

Nayebosadri A, Ji JY. Endothelial nuclear lamina is not required for glucocorticoid receptor nuclear import but does affect receptor-mediated transcription activation. Am J Physiol Cell Physiol 305: C309 -C322, 2013. First published May 22, 2013; doi:10.1152/ajpcell.00293.2012The lamina serves to maintain the nuclear structure and stiffness while acting as a scaffold for heterochromatin and many transcriptional proteins. Its role in endothelial mechanotransduction, specifically how nuclear mechanics impact gene regulation under shear stress, is not fully understood. In this study, we successfully silenced lamin A/C in bovine aortic endothelial cells to determine its role in both glucocorticoid receptor (GR) nuclear translocation and glucocorticoid response element (GRE) transcriptional activation in response to dexamethasone and shear stress. Nuclear translocation of GR, an antiinflammatory nuclear receptor, in response to dexamethasone or shear stress (5, 10, and 25 dyn/cm 2 ) was observed via time-lapse cell imaging and quantified using a Bayesian image analysis algorithm. Transcriptional activity of the GRE promoter was assessed using a dual-luciferase reporter plasmid. We found no dependence on nuclear lamina for GR translocation from the cytoplasm into the nucleus. However, the absence of lamin A/C led to significantly increased expression of luciferase under dexamethasone and shear stress induction as well as changes in histone protein function. PCR results for NF-B inhibitor alpha (NF-BIA) and dual specificity phosphatase 1 (DUSP1) genes further supported our luciferase data with increased expression in the absence of lamin. Our results suggest that absence of lamin A/C does not hinder passage of GR into the nucleus, but nuclear lamina is important to properly regulate GRE transcription. Nuclear lamina, rather than histone deacetylase (HDAC), is a more significant mediator of shear stress-induced transcriptional activity, while dexamethasone-initiated transcription is more HDAC dependent. Our findings provide more insights into the molecular pathways involved in nuclear mechanotransduction. glucocorticoid receptor; nuclear lamina; shear stress; dexamethasone; endothelial cells CARDIOVASCULAR DISEASES SUCH as atherosclerosis are the leading cause of death in the United States. The initiation and development of atherosclerosis are typically associated with endothelial dysfunction (20,51). Endothelial cells at the lumen of blood vessels are continuously exposed to both hemodynamic shear stress and cyclic stretch due to blood flow (16). The distinct local hemodynamic profiles influence endothelial behaviors in the vasculature. Atherosclerotic lesions tend to develop at regions of disturbed flow and low shear stress at bifurcation sites and curvatures, while regions with uniform high laminar shear stress are protected (10,48,66). Endothelial responses to shear stress can be categorized as immediate [rapid release of nitric oxide (NO); (49)] as well as short-term

Phenotypic diversity and glucocorticoid sensitivity in patients with familial partial lipodystrophy type 2

“…More studies, evaluating visceral and peripheral adipose tissues, are necessary to clarify the role of tissue‐specific GC resistance in FPLD2 patients. Finally, we cannot rule out the possibility of nuclear lamina per se being important to properly regulate glucocorticoid response element (GRE) transcription in humans, a mechanism previously reported in bovine aortic endothelial cells …”

Mechanosensing and Mechanoregulation of Endothelial Cell Functions

“…Knockdown of Lamin A reduces sheardependent nuclear translocation of glucocorticoid receptor. Furthermore, shear stress increased HDAC and HAT in control, but not in Lamin A knowndown, suggesting a role for nuclear lamina in regulating chromatin state (273). Modeling studies also suggest that nuclear morphology is crucial for stem cell fate determination, since diffusivity of transcription factors is thought to be a function of local volumetric strain.…”

New Insights in Glucocorticoid Receptor Signaling—More Than Just a Ligand-Binding Receptor

“…Sodium arsenite and dinitrophenol ( 31 ), some conditions such as elevated pH and temperature, and shear stress ( 111 ) can induce GR nuclear translocation in a ligand-independent manner ( 31 , 111 , 112 ) (Figure 1 : 1). Interestingly, GR nuclear translocation induced by shear stress does not depend on ligands or intact cytoskeleton ( 113 ), but it is related to the nuclear lamina ( 114 ).…”