Nuclear Factor-KB Interacts Functionally with the Platelet-derived Growth Factor B-Chain Shear-Stress Response Element in Vascular Endothelial Cells Exposed to Fluid Shear Stress

Hemodynamic forces, such as fluid shear stress, that act on the endothelial lining of the cardiovascular system can modulate the expression of an expanding number of genes crucial for homeostasis and the pathogenesis of vascular disease. A 6-bp core element (5'-GAGACC-3'), defined previously as a shear-stress response element is present in the promoters of many genes, including the PDGF B-chain, whose expression is modulated by shear stress. The identity of the nuclear protein(s) binding to this element has not yet been elucidated. Using electrophoretic mobility shift assays and in vitro DNase I footprinting, we demonstrate that nuclear factor-cB p5O-p65 heterodimers, which accumulate in the nuclei of cultured vascular endothelial cells exposed to fluid shear stress, bind to the PDGF-B shear-stress response element in a specific manner. Mutation of this binding motif abrogated its interaction with p5O-p65 and abolished the ability of the promoter to mediate increased gene expression in endothelial cells exposed to shear stress. Transient cotransfection studies indicate that p50-p65 is able to activate PDGF-B shear-stress response ele-ment-dependent reporter gene expression in these cells. These findings thus implicate nuclear factor-cB in the transactivation of an endothelial gene responding to a defined fluid mechanical force. (J. Clin. Invest. 1995.96:1169-1175.)

[1]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[2]  T. Maniatis,et al.  Transcriptional regulation of endothelial cell adhesion molecules: NF‐κB and cytokine‐inducible enhancers , 1995, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[3]  N. Resnick,et al.  Hemodynamic forces are complex regulators of endothelial gene expression , 1995, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[4]  T. Maniatis,et al.  Endothelial interferon regulatory factor 1 cooperates with NF-kappa B as a transcriptional activator of vascular cell adhesion molecule 1 , 1995, Molecular and cellular biology.

[5]  T. Maniatis,et al.  A striking similarity in the organization of the E-selectin and beta interferon gene promoters , 1994, Molecular and cellular biology.

[6]  C. Scheidereit,et al.  Activation of NF‐kappa B in vivo is regulated by multiple phosphorylations. , 1994, The EMBO journal.

[7]  G. Nabel,et al.  An interaction between the DNA-binding domains of RelA(p65) and Sp1 mediates human immunodeficiency virus gene activation , 1994, Molecular and cellular biology.

[8]  D. Bonthron,et al.  Novel cis-acting elements in the human platelet-derived growth factor B-chain core promoter that mediate gene expression in cultured vascular endothelial cells. , 1994, The Journal of biological chemistry.

[9]  J. Ando,et al.  Shear stress inhibits adhesion of cultured mouse endothelial cells to lymphocytes by downregulating VCAM-1 expression. , 1994, The American journal of physiology.

[10]  J. Whelan,et al.  Cooperativity between two NF-kappa B complexes, mediated by high-mobility-group protein I(Y), is essential for cytokine-induced expression of the E-selectin promoter , 1994 .

[11]  M. Gimbrone,et al.  Shear stress selectively upregulates intercellular adhesion molecule-1 expression in cultured human vascular endothelial cells. , 1994, The Journal of clinical investigation.

[12]  J. Pober,et al.  cAMP and tumor necrosis factor competitively regulate transcriptional activation through and nuclear factor binding to the cAMP-responsive element/activating transcription factor element of the endothelial leukocyte adhesion molecule-1 (E-selectin) promoter. , 1994, The Journal of biological chemistry.

[13]  A. Deisseroth,et al.  Interaction of nuclear proteins with an AP‐1/CRE‐like promoter sequence in the human TNF‐α gene , 1994 .

[14]  S Chien,et al.  Fluid shear stress induces a biphasic response of human monocyte chemotactic protein 1 gene expression in vascular endothelium. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[15]  J. Whelan,et al.  Cyclic AMP-independent ATF family members interact with NF-kappa B and function in the activation of the E-selectin promoter in response to cytokines , 1993, Molecular and cellular biology.

[16]  S. Diamond,et al.  Fluid shear stress induces synthesis and nuclear localization of c-fos in cultured human endothelial cells. , 1993, Biochemical and biophysical research communications.

[17]  W. Greene,et al.  Cross‐coupling of the NF‐kappa B p65 and Fos/Jun transcription factors produces potentiated biological function. , 1993, The EMBO journal.

[18]  Y. Yazaki,et al.  Disruption of cytoskeletal structures mediates shear stress-induced endothelin-1 gene expression in cultured porcine aortic endothelial cells. , 1993, The Journal of clinical investigation.

[19]  T. Maniatis,et al.  Mechanisms of transcriptional synergism between distinct virus-inducible enhancer elements , 1993, Cell.

[20]  G. May,et al.  An EMSA-based method for determining the molecular weight of a protein--DNA complex. , 1993, Nucleic acids research.

[21]  S. Izumo,et al.  Regulation of endothelin 1 gene by fluid shear stress is transcriptionally mediated and independent of protein kinase C and cAMP. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[22]  T. Collins.  Endothelial nuclear factor-kappa B and the initiation of the atherosclerotic lesion. , 1993, Laboratory investigation; a journal of technical methods and pathology.

[23]  R. Ross.  The pathogenesis of atherosclerosis: a perspective for the 1990s , 1993, Nature.

[24]  P. Davies,et al.  Mechanical stress mechanisms and the cell. An endothelial paradigm. , 1993, Circulation research.

[25]  J. Frangos,et al.  Pulsatile and steady flow induces c‐fos expression in human endothelial cells , 1993, Journal of cellular physiology.

[26]  S. Schwartz,et al.  Pharmacology of smooth muscle cell replication. , 1992, Hypertension.

[27]  M. U. Nollert,et al.  Regulation of Genetic Expression in Shear Stress–stimulated Endothelial Cells a , 1992, Annals of the New York Academy of Sciences.

[28]  G. Nolan,et al.  Independent modes of transcriptional activation by the p50 and p65 subunits of NF-kappa B. , 1992, Genes & development.

[29]  P. Baeuerle.  The inducible transcription activator NF-κB: regulation by distinct protein subunits , 1991 .

[30]  G. May,et al.  Purification and characterization of Ku-2, an octamer-binding protein related to the autoantigen Ku. , 1991, The Journal of biological chemistry.

[31]  D. Baltimore,et al.  Activation in vitro of NF-κB" by phosphorylation of its inhibitor IκB" , 1990, Nature.

[32]  L V McIntire,et al.  Fluid flow stimulates tissue plasminogen activator secretion by cultured human endothelial cells. , 1989, Science.

[33]  L V McIntire,et al.  Flow effects on prostacyclin production by cultured human endothelial cells. , 1985, Science.

[34]  S. Ruben,et al.  Selection of Optimal KB/Rel DNA-Binding Motifs: Interaction of Both Subunits of NF-iB with DNA Is Required for Transcriptional Activation , 2003 .

[35]  Cytoskeletal Control of Gene Expression : Depolymerization of Microtubules Activates NF-xB , 2002 .

[36]  C F Dewey,et al.  Platelet-derived growth factor B chain promoter contains a cis-acting fluid shear-stress-responsive element. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[37]  M. Grilli,et al.  NF-kappa B and Rel: participants in a multiform transcriptional regulatory system. , 1993, International review of cytology.