Signaling mechanisms underlying strain-dependent brain natriuretic peptide gene transcription

Activation of brain natriuretic peptide (BNP) gene promoter activity represents one of the earliest and most reliable markers of ventricular cardiac myocyte hypertrophy. We recently demonstrated that mechanical strain increases immunoreactive BNP secretion, steady-state BNP mRNA levels and BNP gene transcriptional activity in neonatal rat myocyte cultures. We have also shown that strain-dependent BNP gene transcription is critically dependent on the functional integrity of a number of integrins (specfically β1, β3, and αvβ5 integrins) present on the surface of cardiac myocytes. When used alone, each of these antibodies resulted in a significant reduction in strain-dependent activation of a transfected hBNP-luciferase reporter and inhibition of a number of signaling pathways that have been linked to stimulation of this reporter (e.g., extracellular signal regulated kinase and c-Jun amino terminal kinase). The present study shows that combinations of these antibodies resulted in further reductions in hBNP gene promoter activity and inhibition of the relevant signaling cascades. These studies provide further support for the importance of integrin-matrix interactions in promoting strain-dependent changes in cardiac myocyte gene transcription.Key words: mechanical strain, brain natriuretic peptide, integrins, mitogen-activated protein kinase, cardiac myocyte.

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Integrin Dependence of Brain Natriuretic Peptide Gene Promoter Activation by Mechanical Strain

Journal of Biological Chemistry ◽

10.1074/jbc.m001660200 ◽

2000 ◽

Vol 275 (27) ◽

pp. 20355-20360 ◽

Cited By ~ 32

Author(s):

Faquan Liang ◽

Amha Atakilit ◽

David G. Gardner

Keyword(s):

Brain Natriuretic Peptide ◽

Natriuretic Peptide ◽

Mechanical Strain ◽

Gene Promoter ◽

Promoter Activation ◽

Peptide Gene

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Phospholipase A 2 Metabolites Mediate Endothelin Stimulation of the Human Brain Natriuretic Peptide Promoter

Hypertension ◽

10.1161/hyp.36.suppl_1.721-b ◽

2000 ◽

Vol 36 (suppl_1) ◽

pp. 721-721

Author(s):

Quan He

Keyword(s):

Gene Expression ◽

Brain Natriuretic Peptide ◽

Natriuretic Peptide ◽

Promoter Activity ◽

Luciferase Activity ◽

Phospholipase A ◽

Luciferase Reporter ◽

Luciferase Reporter Gene ◽

P450 Monooxygenase ◽

Stimulation Of

P155 Brain natriuretic peptide (BNP) gene expression accompanies cardiac hypertrophy and heart failure. The vasoconstrictor endothelin-1 (ET)may be involved in the development of these diseases. ET has also been shown to activate phospholipase A 2 (PLA 2 ). Thus we studied whether ET and PLA 2 metabolites regulate BNP gene expression. The hBNP promoter (-1818 to + 100) coupled to a luciferase reporter gene was transferred into neonatal ventricular myocytes (NVM),and luciferase activity was measured as an index of promoter activity. ET (10 -7 M)induced BNP mRNA in NVM as assessed by Northern blot. It also stimulated the hBNP promoter 4-fold vs control, an effect completely inhibited by actinomycin D. To test the involvement of different PLA 2 isoforms, transfected cells were treated with the Ca ++ -independent PLA 2 (iPLA 2 )inhibitor bromoenol lactone (BEL), the cytosolic PLA 2 inhibitor methyl arachidonyl fluorophosphonate, or the secretory PLA 2 inhibitor ONO-RS-082 prior to stimulation with ET. Only the iPLA 2 inhibitor BEL prevented ET-stimulated hBNP promoter activity. The PLA 2 metabolite lysophosphatidic acid (LPA) also activated the hBNP promoter (2.2-fold; n = 3), but lysophosphatidylcholine did not. To test whether arachidonic acid metabolites are involved in ET’s effect, cells were pretreated with either a lipoxygenase (LO), cyclooxygenase, or p450 monooxygenase inhibitor. Only the LO inhibitor baicalein prevented ET stimulation of the hBNP promoter. Finally, we studied the involvement of cis elements in ET-stimulated hBNP promoter activity. Deletion of BNP promoter sequences from -1818 to -408 and from -408 to -40 reduced ET’s effect by 54% and 78%, respectively. Moreover, ET-stimulated luciferase activity was reduced by 53% when the GATA element (at position -85 relative to the start site of transcription) was mutated. These data suggest that: 1) ET activates the hBNP promoter through a transcriptional mechanism; 2) LPA, perhaps generated by a BEL-sensitive iPLA 2 , is involved in ET’s effect; 3) a LO pathway may also mediate ET signaling; and 4) ET regulation of the hBNP promoter targets both distal and proximal cis elements, including GATA.

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Isoproterenol and cAMP regulation of the human brain natriuretic peptide gene involves Src and Rac

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.2000.278.6.e1115 ◽

2000 ◽

Vol 278 (6) ◽

pp. E1115-E1123 ◽

Cited By ~ 27

Author(s):

Quan He ◽

Guiyun Wu ◽

Margot C. Lapointe

Keyword(s):

Brain Natriuretic Peptide ◽

Natriuretic Peptide ◽

Promoter Activity ◽

Kinase Inhibitor ◽

Regulatory Elements ◽

Dominant Negative ◽

Luciferase Reporter ◽

Dominant Negative Mutant ◽

Luciferase Reporter Gene ◽

Src Tyrosine Kinase

Brain natriuretic peptide (BNP) gene expression and chronic activation of the sympathetic nervous system are characteristics of the development of heart failure. We studied the role of the β-adrenergic signaling pathway in regulation of the human BNP (hBNP) promoter. An hBNP promoter (−1818 to +100) coupled to a luciferase reporter gene was transferred into neonatal cardiac myocytes, and luciferase activity was measured as an index of promoter activity. Isoproterenol (ISO), forskolin, and cAMP stimulated the promoter, and the β2-antagonist ICI 118,551 abrogated the effect of ISO. In contrast, the protein kinase A (PKA) inhibitor H-89 failed to block the action of cAMP and ISO. Pertussis toxin (PT), which inactivates Gαi, inhibited ISO- and cAMP-stimulated hBNP promoter activity. The Src tyrosine kinase inhibitor PP1 and a dominant-negative mutant of the small G protein Rac also abolished the effect of ISO and cAMP. Finally, we studied the involvement of M-CAT-like binding sites in basal and inducible regulation of the hBNP promoter. Mutation of these elements decreased basal and cAMP-induced activity. These data suggest that β-adrenergic regulation of hBNP is PKA independent, involves a Gαi-activated pathway, and targets regulatory elements in the proximal BNP promoter.

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Regulation of the human brain natriuretic peptide gene by GATA-4

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00274.2001 ◽

2002 ◽

Vol 283 (1) ◽

pp. E50-E57 ◽

Cited By ~ 26

Author(s):

Quan He ◽

Mariela Mendez ◽

Margot C. LaPointe

Keyword(s):

Brain Natriuretic Peptide ◽

Natriuretic Peptide ◽

Promoter Activity ◽

Nuclear Protein ◽

Ventricular Myocytes ◽

Proximal Promoter ◽

Luciferase Reporter ◽

Luciferase Reporter Gene ◽

Mobility Shift ◽

Specific Expression

Brain natriuretic peptide (BNP) is a cardiac hormone constitutively expressed in the adult heart. We previously showed that the human BNP (hBNP) proximal promoter region from −127 to −40 confers myocyte-specific expression. The proximal hBNP promoter contains several putative cis elements. Here we tested whether the proximal GATA element plays a role in basal and inducible regulation of the hBNP promoter. The hBNP promoter was coupled to a luciferase reporter gene (1818hBNPLuc) and transferred into neonatal ventricular myocytes (NVM), and luciferase activity was measured as an index of hBNP promoter activity. Mutation of the putative GATA element at −85 of the hBNP promoter [1818(mGATA)hBNPLuc] reduced activity by 97%. To study transactivation of the hBNP promoter, we co-transfected 1818hBNPLuc with the GATA-4 expression vector. GATA-4 activated 1818hBNPLuc, and this effect was eliminated by mutation of the proximal GATA element. Electrophoretic mobility shift assay showed that an oligonucleotide containing the hBNP GATA motif bound to cardiomyocyte nuclear protein, which was competed for by a consensus GATA oligonucleotide but not a mutated hBNP GATA element. The β-adrenergic agonist isoproterenol and its second messenger cAMP stimulated hBNP promoter activity and binding of nuclear protein to the proximal GATA element. Thus the GATA element in the proximal hBNP promoter is involved in both basal and inducible transcriptional regulation in cardiac myocytes.

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Focal Adhesion Kinase and p130Cas Mediate Both Sarcomeric Organization and Activation of Genes Associated with Cardiac Myocyte Hypertrophy

Molecular Biology of the Cell ◽

10.1091/mbc.12.8.2290 ◽

2001 ◽

Vol 12 (8) ◽

pp. 2290-2307 ◽

Cited By ~ 67

Author(s):

Branka Kovac̆ic̆-Milivojević ◽

Frederick Roediger ◽

Eduardo A.C. Almeida ◽

Caroline H. Damsky ◽

David G. Gardner ◽

...

Keyword(s):

Gene Expression ◽

Focal Adhesion Kinase ◽

Cardiac Myocytes ◽

Natriuretic Peptide ◽

Focal Adhesion ◽

Promoter Activity ◽

Cardiac Myocyte ◽

Gene Promoter ◽

Myocyte Hypertrophy ◽

Cardiac Myocyte Hypertrophy

Hypertrophic terminally differentiated cardiac myocytes show increased sarcomeric organization and altered gene expression. Previously, we established a role for the nonreceptor tyrosine kinase Src in signaling cardiac myocyte hypertrophy. Here we report evidence that p130Cas (Cas) and focal adhesion kinase (FAK) regulate this process. In neonatal cardiac myocytes, tyrosine phosphorylation of Cas and FAK increased upon endothelin (ET) stimulation. FAK, Cas, and paxillin were localized in sarcomeric Z-lines, suggesting that the Z-line is an important signaling locus in these cells. Cas, alone or in cooperation with Src, modulated basal and ET-stimulated atrial natriuretic peptide (ANP) gene promoter activity, a marker of cardiac hypertrophy. Expression of the C-terminal focal adhesion-targeting domain of FAK interfered with localization of endogenous FAK to Z-lines. Expression of the Cas-binding proline-rich region 1 of FAK hindered association of Cas with FAK and impaired the structural stability of sarcomeres. Collectively, these results suggest that interaction of Cas with FAK, together with their localization to Z-lines, is critical to assembly of sarcomeric units in cardiac myocytes in culture. Moreover, expression of the focal adhesion-targeting and/or the Cas-binding proline-rich regions of FAK inhibited ANP promoter activity and suppressed ET-induced ANP and brain natriuretic peptide gene expression. In summary, assembly of signaling complexes that include the focal adhesion proteins Cas, FAK, and paxillin at Z-lines in the cardiac myocyte may regulate, either directly or indirectly, both cytoskeletal organization and gene expression associated with cardiac myocyte hypertrophy.

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PGE2stimulates human brain natriuretic peptide expression via EP4and p42/44 MAPK

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00904.2005 ◽

2006 ◽

Vol 290 (5) ◽

pp. H1740-H1746 ◽

Cited By ~ 22

Author(s):

Jian-Yong Qian ◽

Alicia Leung ◽

Pamela Harding ◽

Margot C. LaPointe

Keyword(s):

Protein Synthesis ◽

Brain Natriuretic Peptide ◽

Natriuretic Peptide ◽

Promoter Activity ◽

Small Gtpases ◽

Dominant Negative ◽

Luciferase Reporter ◽

Dominant Negative Mutant ◽

Prostaglandin E ◽

Stimulation Of

Brain natriuretic peptide (BNP) produced by cardiac myocytes has antifibrotic and antigrowth properties and is a marker of cardiac hypertrophy. We previously showed that prostaglandin E2(PGE2) is the main prostaglandin produced in myocytes treated with proinflammatory stimuli and stimulates protein synthesis by binding to its EP4receptor. We hypothesized that PGE2, acting through EP4, also regulates BNP gene expression. We transfected neonatal ventricular myocytes with a plasmid encoding the human BNP (hBNP) promoter driving expression of a luciferase reporter gene. PGE2increased hBNP promoter activity 3.5-fold. An EP4antagonist reduced the stimulatory effect of PGE2but not an EP1antagonist. Because EP4signaling can involve adenylate cyclase, cAMP, and protein kinase A (PKA), we tested the effect of H-89, a PKA inhibitor, on PGE2stimulation of the hBNP promoter. H-89 at 5 μM decreased PGE2stimulation of BNP promoter activity by 100%. Because p42/44 MAPK mediates the effect of PGE2on protein synthesis, we also examined the role of MAPKs in the regulation of BNP promoter activity. PGE2stimulation of the hBNP promoter was inhibited by a MEK1/2 inhibitor and a dominant-negative mutant of Raf, indicating that p42/44 MAPK was involved. In contrast, neither a p38 MAPK inhibitor nor a JNK inhibitor reduced the stimulatory effect of PGE2. Involvement of small GTPases was also studied. Dominant-negative Rap inhibited PGE2stimulation of the hBNP promoter, but dominant-negative Ras did not. We concluded that PGE2stimulates the BNP promoter mainly via EP4, PKA, Rap, and p42/44 MAPK.

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