title: Endothelial nuclear lamina is not required for glucocorticoid receptor nuclear import but does affect receptor-mediated transcription activation.
abstract: The 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 anti-inflammatory 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-κB inhibitor alpha (NF-κBIA) 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.
authors.affiliation: Department of Biomedical Engineering, Indiana University Purdue University Indianapolis, Indiana University Purdue University Indianapolis, Indianapolis, Indiana, USA.
authors.forename: Arman
authors.initials: A
authors.lastname: Nayebosadri
authors.forename: Julie Y
authors.initials: JY
authors.lastname: Ji
mesh_terms: D021581:Active Transport, Cell Nucleus; D000818:Animals; D001011:Aorta / Q000166:cytology; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D003907:Dexamethasone / Q000494:pharmacology; D054638:Dual Specificity Phosphatase 1 / Q000096:biosynthesis; D042783:Endothelial Cells / Q000378:metabolism; D005786:Gene Expression Regulation; D006570:Heterochromatin; D006655:Histone Deacetylases / Q000378:metabolism; D020672:I-kappa B Proteins / Q000096:biosynthesis; D034904:Lamin Type A / Q000235:genetics; D008156:Luciferases / Q000096:biosynthesis; D040542:Mechanotransduction, Cellular; D051379:Mice; D018345:Mice, Knockout; D000072000:NF-KappaB Inhibitor alpha; D034881:Nuclear Lamina / Q000378:metabolism*; D034622:RNA Interference; D034741:RNA, Small Interfering; D011965:Receptors, Glucocorticoid / Q000378:metabolism*; D020218:Response Elements / Q000235:genetics; D013312:Stress, Physiological; D015533:Transcriptional Activation
chemical_list: D006570:Heterochromatin; D020672:I-kappa B Proteins; D034904:Lamin Type A; C000606205:Nfkbia protein, mouse; D034741:RNA, Small Interfering; D011965:Receptors, Glucocorticoid; D000072000:NF-KappaB Inhibitor alpha; D003907:Dexamethasone; D008156:Luciferases; D054638:Dual Specificity Phosphatase 1; C517020:Dusp1 protein, mouse; D006655:Histone Deacetylases
keywords: dexamethasone; endothelial cells; glucocorticoid receptor; nuclear lamina; shear stress