Endothelial Yin Yang 1 Phosphorylation at S118 Induces Atherosclerosis Under Flow

Rationale: Disturbed flow occurring in arterial branches and curvatures induces vascular endothelial cell (EC) dysfunction and atherosclerosis. We postulated that disturbed flow plays important roles in modulating phosphoprotein expression profiles to regulate endothelial functions and atherogenesis. Objective: The goal of this study is to discover novel site-specific phosphorylation alterations induced by disturbed flow in ECs to contribute to atherosclerosis. Methods and Results: Quantitative phosphoproteomics analysis of ECs exposed to disturbed flow with low and oscillatory shear stress (OS, 0.5plusminus4 dynes/cm 2 ) vs. pulsatile flow with high shear stress (PS, 124plusminus dynes/cm 2 ) revealed that OS induces serine (S)118 phosphorylation of Yin Yang 1 (phospho-YY1 S118 ) in ECs. Elevated phospho-YY1 S118 level in ECs was further confirmed to be present in the disturbed flow regions in experimental animals and human atherosclerotic arteries. This disturbed flow-induced EC phospho-YY1 S118 is mediated by casein kinase 2α (CK2α) through its direct interaction with YY1. Yeast two-hybrid library screening and in situ proximity ligation assays demonstrate that phospho-YY1 S118 directly binds zinc finger with KRAB and SCAN domains 4 (ZKSCAN4) to induce promoter activity and gene expression of human double minute 2 (HDM2), which consequently induces EC proliferation through down-regulations of p53 and p21 CIP1 . Administration of apolipoprotein E-deficient (ApoE -/- ) mice with CK2-specific inhibitor tetrabromocinnamic acid or atorvastatin inhibits atherosclerosis formation through down-regulations of EC phospho-YY1 S118 and HDM2. Generation of novel transgenic mice bearing EC-specific overexpression of S118-non-phosphorylatable mutant of YY1 in ApoE -/- mice confirms the critical role of phospho-YY1 S118 in promoting atherosclerosis through EC HDM2. Conclusions: Our findings provide new insights into the mechanisms by which disturbed flow induces endothelial phospho-YY1 S118 to promote atherosclerosis, thus indicating phospho-YY1 S118 as a potential molecular target for atherosclerosis treatment.

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Abstract 363: Lim Domain Only 4 Is a Shear-Sensitive Protein, Playing a Critical Role in Endothelial Inflammation and Atherosclerosis

Arteriosclerosis Thrombosis and Vascular Biology ◽

10.1161/atvb.32.suppl_1.a363 ◽

2012 ◽

Vol 32 (suppl_1) ◽

Author(s):

Wakako Takabe ◽

Chih-Wen Ni ◽

Dong Ju Son ◽

Noah Alberts-Grill ◽

Hanjoong Jo

Keyword(s):

Endothelial Cells ◽

Shear Stress ◽

Critical Role ◽

Oscillatory Shear ◽

Lim Domain ◽

Disturbed Flow ◽

Endothelial Inflammation ◽

And Migration ◽

Oscillatory Shear Stress ◽

Stable Flow

Recently, we have shown that disturbed flow, characterized by low and oscillatory shear stress, caused by a partial ligation of mouse left carotid artery (LCA) rapidly induces atherosclerosis. Using the partial ligation model and genome-wide microarray study with aortic endothelial RNAs obtained directly from the flow-disturbed carotid arteries, we previously identified mechanosensitive genes in mouse endothelial RNA including LIM domain only 4 ( lmo4 ). Here we report that LMO4 is a shear-sensitive protein that regulates endothelial inflammation. Lmo4 was up-regulated by disturbed flow in mouse LCA compared to the contralateral right CA (RCA) exposed to stable flow. At protein levels, LMO4 expression was significantly higher not only in LCA in our surgical model but also in the lesser curvature (flow-disturbed and athero-prone region of mouse aortic arch) compared to the greater curvature (stable-flow and ather-protected region). In addition, immunohistochemical staining of LMO4 in human coronary arteries revealed that its expression is detectable only in intimal endothelial cells, but not in medial cells. While LMO4 is known as a potential oncogene and associated with growth, migration and invasion of breast cancer cells, its role in cardiovascular system is not known to our knowledge. We tested a hypothesis that LMO4 is a mechanosensitive gene and plays a critical role in regulation of endothelial cell biology. LMO4 protein expression was robustly induced by oscillatory shear stress (OS) compared to laminar shear (LS) in human umbilical vein endothelial cells (HUVEC). Treatment of HUVEC with siRNA against LMO4 significantly inhibited OS-induced inflammation and migration, but not apoptosis and cell cycle progression. Further, LMO4 siRNA treatment significantly blunted expression of VCAM-1 and interleukin-8 induced by OS in endothelial cells. These results suggest that LMO4 is a shear-induced gene that plays a critical role in OS-induced endothelial inflammation and migration, and potentially in atherosclerosis.

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Astrocyte-specific deletion of the transcription factor Yin Yang 1 in murine substantia nigra mitigates manganese-induced dopaminergic neurotoxicity

Journal of Biological Chemistry ◽

10.1074/jbc.ra120.015552 ◽

2020 ◽

Vol 295 (46) ◽

pp. 15662-15676 ◽

Cited By ~ 1

Author(s):

Edward Pajarillo ◽

James Johnson ◽

Asha Rizor ◽

Ivan Nyarko-Danquah ◽

Getinet Adinew ◽

...

Keyword(s):

Transcription Factor ◽

Substantia Nigra ◽

Critical Role ◽

Conditional Knockout ◽

Motor Functions ◽

Mn Toxicity ◽

Protein Levels ◽

Yin Yang 1 ◽

Yin Yang

Manganese (Mn)-induced neurotoxicity resembles Parkinson's disease (PD), but the mechanisms underpinning its effects remain unknown. Mn dysregulates astrocytic glutamate transporters, GLT-1 and GLAST, and dopaminergic function, including tyrosine hydroxylase (TH). Our previous in vitro studies have shown that Mn repressed GLAST and GLT-1 via activation of transcription factor Yin Yang 1 (YY1). Here, we investigated if in vivo astrocytic YY1 deletion mitigates Mn-induced dopaminergic neurotoxicity, attenuating Mn-induced reduction in GLAST/GLT-1 expression in murine substantia nigra (SN). AAV5-GFAP-Cre-GFP particles were infused into the SN of 8-week–old YY1flox/flox mice to generate a region-specific astrocytic YY1 conditional knockout (cKO) mouse model. 3 weeks after adeno-associated viral (AAV) infusion, mice were exposed to 330 μg of Mn (MnCl2 30 mg/kg, intranasal instillation, daily) for 3 weeks. After Mn exposure, motor functions were determined in open-field and rotarod tests, followed by Western blotting, quantitative PCR, and immunohistochemistry to assess YY1, TH, GLAST, and GLT-1 levels. Infusion of AAV5-GFAP-Cre-GFP vectors into the SN resulted in region-specific astrocytic YY1 deletion and attenuation of Mn-induced impairment of motor functions, reduction of TH-expressing cells in SN, and TH mRNA/protein levels in midbrain/striatum. Astrocytic YY1 deletion also attenuated the Mn-induced decrease in GLAST/GLT-1 mRNA/protein levels in midbrain. Moreover, YY1 deletion abrogated its interaction with histone deacetylases in astrocytes. These results indicate that astrocytic YY1 plays a critical role in Mn-induced neurotoxicity in vivo, at least in part, by reducing astrocytic GLAST/GLT-1. Thus, YY1 might be a potential target for treatment of Mn toxicity and other neurological disorders associated with dysregulation of GLAST/GLT-1.

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The Adaptor Protein Nck1, but not Nck2, Mediates Shear Stress-Induced Endothelial Permeability

10.1101/651687 ◽

2019 ◽

Author(s):

Mabruka Alfaidi ◽

Umesh Bhattarai ◽

Elizabeth D Cockerham ◽

A.W. Orr

Keyword(s):

Shear Stress ◽

Vascular Permeability ◽

Pore Formation ◽

Critical Role ◽

Endothelial Permeability ◽

Adaptor Protein ◽

Adaptor Proteins ◽

Disturbed Flow

AbstractAlteration in hemodynamic shear stress at atheroprone sites promotes endothelial paracellular pore formation and permeability. Previously, we have reported that a peptide inhibitor to Nck prevented shear stress-induced p21 activated kinase (PAK) activation and endothelial permeability. However, the specificity of this peptide is unclear, and the role of individual Nck isoforms remain unknown. Here, we show that genetic deletion of Nck1/2 adaptor proteins significantly ameliorates shear stress induced permeability, and selective isoform depletion suggests distinct signaling mechanisms. Only Nck1 deletion significantly reduces flow-induced paracellular pore formation and permeability, whereas Nck2 depletion has no significant effects. Additionally, Nck1 reexpression, but not Nck2, restores shear stress-induced permeability in Nck1/2 knockout cells, confirming the non-compensating roles. In vivo, using the partial carotid ligation model of disturbed flow, Nck1 knockout prevented the increase in vascular permeability, as assessed by both Evans blue extravasation and leakage of plasma fibrinogen into the vessel wall. Domain swap experiments mixing SH2 (phosphotyrosine binding) and SH3 (proline rich binding) domains between Nck1 and Nck2 showed a dispensable role for SH2 domains but a critical role for the Nck1 SH3 domains in rescuing shear stress-induced endothelial permeability. Consistent with this, both Nck1 and Nck2 bind to PECAM-1 (SH2 dependent) in response to shear stress, but only Nck1 ablation interferes with shear stress-induced PAK2 activation (SH3 dependent). This work provides the first evidence that Nck1 and Nck2 play distinct roles in flow-induced vascular permeability.New and NoteworthyThe present study shows a specific role for Nck1 in endothelial permeability in response to shear stress. Using in vitro and in vivo models, we demonstrate improvement in endothelial barrier integrity in cells subjected to disturbed flow only following Nck1 but not Nck2 deletion. Selective Nck1 inhibition may limit endothelial permeability at sites of disturbed flow to reduce atherosclerosis without affecting angiogenesis, which requires both Nck1 and Nck2 inhibition.

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Canine Angiostrongylus vasorum-Induced Early Innate Immune Reactions Based on NETs Formation and Canine Vascular Endothelial Cell Activation In Vitro

Biology ◽

10.3390/biology10050427 ◽

2021 ◽

Vol 10 (5) ◽

pp. 427

Author(s):

Daniela Grob ◽

Iván Conejeros ◽

Sara López-Osorio ◽

Zahady D. Velásquez ◽

Lisa Segeritz ◽

...

Keyword(s):

Endothelial Cells ◽

Cell Activation ◽

Expression Profiles ◽

Critical Role ◽

Vascular Endothelial Cell ◽

Angiostrongylus Vasorum ◽

Polymorphonuclear Neutrophils ◽

Endothelial Cell Activation ◽

Immune Reactions

Due to its localization in the canine blood stream, Angiostrongylus vasorum is exposed to circulating polymorphonuclear neutrophils (PMN) and the endothelial cells of vessels. NETs release of canine PMN exposed to A. vasorum infective stages (third stage larvae, L3) and early pro-inflammatory immune reactions of primary canine aortic endothelial cells (CAEC) stimulated with A. vasorum L3-derived soluble antigens (AvAg) were analyzed. Expression profiles of the pro-inflammatory adhesion molecules ICAM-1, VCAM-1, P-selectin and E-selectin were analyzed in AvAg-stimulated CAEC. Immunofluorescence analyses demonstrated that motile A. vasorum L3 triggered different NETs phenotypes, with spread NETs (sprNETs) as the most abundant. Scanning electron microscopy confirmed that the co-culture of canine PMN with A. vasorum L3 resulted in significant larval entanglement. Distinct inter-donor variations of P-selectin, E-selectin, ICAM-1 and VCAM-1 gene transcription and protein expression were observed in CAEC isolates which might contribute to the high individual variability of pathological findings in severe canine angiostrongylosis. Even though canine NETs did not result in larval killing, the entanglement of L3 might facilitate further leukocyte attraction to their surface. Since NETs have already been documented as involved in both thrombosis and endothelium damage events, we speculate that A. vasorum-triggered NETs might play a critical role in disease outcome in vivo.

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Myocardial BDNF regulates cardiac bioenergetics through the transcription factor Yin Yang 1

10.1101/2021.01.19.427213 ◽

2021 ◽

Author(s):

Xue Yang ◽

Manling Zhang ◽

Raymond J. Zimmerman ◽

Qin Wang ◽

An-chi Wei ◽

...

Keyword(s):

Heart Failure ◽

Transcription Factor ◽

Critical Role ◽

Mitochondrial Protein ◽

Cardiac Response ◽

Mouse Heart ◽

Bdnf Expression ◽

Yin Yang 1 ◽

Mitochondrial Dna Copy Number ◽

Yin Yang

ABSTRACTSerum Brain-derived Neurotrophic Factor (BDNF) is markedly decreased in heart failure patients. Both BDNF and its receptor, Tropomyosin Related Kinase Receptor (TrkB), are expressed in cardiomyocytes, however the role of myocardial BDNF signaling in cardiac pathophysiology is poorly understood. We found that myocardial BDNF expression was increased in mice with swimming exercise, but decreased in a mouse heart failure model. Cardiac-specific TrkB knockout (cTrkB KO) mice displayed a blunted adaptive cardiac response to exercise, with attenuated upregulation of transcription factor networks controlling mitochondrial biogenesis/metabolism, including Peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC-1α). In response to pathological stress (transaortic constriction, TAC), cTrkB KO mice showed an exacerbated heart failure progression. The expression of PGC-1α and other metabolic transcription factors were downregulated in cTrkB KO mice exposed to TAC. Consistent with this, mitochondrial DNA copy number and mitochondrial protein abundance was markedly decreased in cTrkB KO mice, resulting in decreased mitochondrial respiratory function. We further unraveled that BDNF induces PGC-1α upregulation and bioenergetics through a novel signaling pathway, the pleiotropic transcription factor Yin Yang 1 (YY1). Taken together, our findings suggest that myocardial BDNF plays a critical role in regulating cellular energetics in the cardiac stress response.

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Role of p38 MAP kinase in endothelial cell alignment induced by fluid shear stress

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.2001.280.1.h189 ◽

2001 ◽

Vol 280 (1) ◽

pp. H189-H197 ◽

Cited By ~ 79

Author(s):

Nobuyoshi Azuma ◽

Nobuyuki Akasaka ◽

Hiroyuki Kito ◽

Masataka Ikeda ◽

Vivian Gahtan ◽

...

Keyword(s):

Shear Stress ◽

Map Kinase ◽

Fluid Shear Stress ◽

Morphological Changes ◽

Critical Role ◽

Specific Inhibitor ◽

Orientation Angle ◽

Fiber Formation ◽

Dominant Negative Mutant ◽

Sb 203580

The p38/mitogen-activated protein (MAP) kinase-activated protein kinase 2 (MAPKAP kinase 2)/heat shock protein (HSP)25/27 pathway is thought to play a critical role in actin dynamics. In the present study, we examined whether p38 was involved in the morphological changes seen in endothelial cells (EC) exposed to shear stress. Cultured bovine aortic EC were subjected to 14 dyn/cm2 laminar steady shear stress. Peak activation of p38, MAPKAP kinase 2, and HSP25 were sixfold at 5 min, sixfold at 5 min, and threefold at 30 min compared with static control, respectively. SB-203580 (1 μM), a specific inhibitor of p38, abolished the activation of MAPKAP kinase 2 and HSP25 as well as EC elongation and alignment in the direction of flow elicited by shear stress. The mean orientation angle of cells subjected to shear without SB-203580, with SB-203580, or static control were 17, 50, and 43°, respectively ( P < 0.05). EC transfected with the dominant negative mutant of p38-α aligned randomly with no stress fiber formation despite exposure to shear stress. These data suggests that the pathway of p38/MAPKAP kinase 2/HSP25/27 is activated in response to shear stress, and this pathway plays an important role in morphological changes induced by shear stress.

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