Cellular Distribution and Function of Soluble Guanylyl Cyclase in Rat Kidney and Liver

Abstract. Soluble guanylyl cyclase (sGC) catalyzes the biosynthesis of cGMP in response to binding of L-arginine-derived nitric oxide (NO). Functionally, the NO-sGC-cGMP signaling pathway in kidney and liver has been associated with regional hemodynamics and the regulation of glomerular parameters. The distribution of the ubiquitous sGC isoform α1β1 sGC was studied with a novel, highly specific antibody against the β1 subunit. In parallel, the presence of mRNA encoding both subunits was investigated by using in situ hybridization and reverse transcription-PCR assays. The NO-induced, sGC-dependent accumulation of cGMP in cytosolic extracts of tissues and cells was measured in vitro. Renal glomerular arterioles, including the renin-producing granular cells, mesangium, and descending vasa recta, as well as cortical and medullary interstitial fibroblasts, expressed sGC. Stimulation of isolated mesangial cells, renal fibroblasts, and hepatic Ito cells with a NO donor resulted in markedly increased cytosolic cGMP levels. This assessment of sGC expression and activity in vascular and interstitial cells of kidney and liver may have implications for understanding the role of local cGMP signaling cascades.

Download Full-text

45 years of cGMP research in Dictyostelium: Understanding the regulation and function of the cGMP pathway for cell movement and chemotaxis

Molecular Biology of the Cell ◽

10.1091/mbc.e21-04-0171 ◽

2021 ◽

pp. mbc.E21-04-0171

Author(s):

Peter J.M. van Haastert ◽

Ineke Keizer-Gunnink ◽

Henderikus Pots ◽

Claudia Ortiz-Mateos ◽

Douwe Veltman ◽

...

Keyword(s):

Signaling Pathway ◽

Guanylyl Cyclase ◽

Cell Movement ◽

Soluble Guanylyl Cyclase ◽

Leading Edge ◽

Actin Binding ◽

Myosin Filament ◽

Cgmp Pathway ◽

Cgmp Signaling ◽

And Function

In Dictyostelium chemoattractants induce a fast cGMP response that mediates myosin filament formation in the rear of the cell. The major cGMP signaling pathway consists of a soluble guanylyl cyclase sGC, a cGMP-stimulated cGMP-specific phosphodiesterase and the cGMP-target protein GbpC. Here we combine published experiments with many unpublished experiments performed in the past 45 years on the regulation and function of the cGMP signaling pathway. The chemoattractants stimulate heterotrimeric Gαβγ and monomeric Ras proteins. A fraction of the soluble guanylyl cyclase sGC binds with high affinity to a limited number of membrane binding site, which is essential for sGC to become activated by Ras and Gα proteins. sGC can also bind to F-actin; binding to branched F-actin in pseudopods enhances basal sGC activity, whereas binding to parallel F-actin in the cortex reduces sGC activity. The cGMP pathway mediates cell polarity by inhibiting the rear: in unstimulated cells by sGC activity in the branched F-actin of pseudopods, in a shallow gradient by stimulated cGMP formation in pseudopods at the leading edge, and during cAMP oscillation to erase the previous polarity and establish a new polarity axis that aligns with the direction of the passing cAMP wave.

Download Full-text

Biphasic roles for soluble guanylyl cyclase (sGC) in platelet activation

Blood ◽

10.1182/blood-2011-03-341107 ◽

2011 ◽

Vol 118 (13) ◽

pp. 3670-3679 ◽

Cited By ~ 45

Author(s):

Guoying Zhang ◽

Binggang Xiang ◽

Anping Dong ◽

Radek C. Skoda ◽

Alan Daugherty ◽

...

Keyword(s):

Platelet Activation ◽

Guanylyl Cyclase ◽

Soluble Guanylyl Cyclase ◽

Whole Body ◽

No Donor ◽

No Donors ◽

Deficient Mice

AbstractNitric oxide (NO) stimulates cGMP synthesis by activating its intracellular receptor, soluble guanylyl cyclase (sGC). It is a currently prevailing concept that No and cGMP inhibits platelet function. However, the data supporting the inhibitory role of NO/sGC/cGMP in platelets have been obtained either in vitro or using whole body gene deletion that affects vessel wall function. Here we have generated mice with sGC gene deleted only in megakaryocytes and platelets. Using the megakaryocyte- and platelet-specific sGC-deficient mice, we identify a stimulatory role of sGC in platelet activation and in thrombosis in vivo. Deletion of sGC in platelets abolished cGMP production induced by either NO donors or platelet agonists, caused a marked defect in aggregation and attenuated secretion in response to low doses of collagen or thrombin. Importantly, megakaryocyte- and platelet-specific sGC deficient mice showed prolonged tail-bleeding times and impaired FeCl3-induced carotid artery thrombosis in vivo. Interestingly, the inhibitory effect of the NO donor SNP on platelet activation was sGC-dependent only at micromolar concentrations, but sGC-independent at millimolar concentrations. Together, our data demonstrate important roles of sGC in stimulating platelet activation and in vivo thrombosis and hemostasis, and sGC-dependent and -independent inhibition of platelets by NO donors.

Download Full-text

Low guanylyl cyclase activity in Weddell seals: implications for peripheral vasoconstriction and perfusion of the brain during diving

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.00283.2018 ◽

2019 ◽

Vol 316 (6) ◽

pp. R704-R715 ◽

Cited By ~ 4

Author(s):

Allyson G. Hindle ◽

Kaitlin N. Allen ◽

Annabelle J. Batten ◽

Luis A. Hückstädt ◽

Jason Turner-Maier ◽

...

Keyword(s):

Guanylyl Cyclase ◽

Soluble Guanylyl Cyclase ◽

Amino Acid Sequences ◽

No Production ◽

Diving Response ◽

No Donor ◽

Weddell Seals ◽

Terrestrial Mammals ◽

And Function ◽

The Brain

Nitric oxide (NO) is a potent vasodilator, which improves perfusion and oxygen delivery during tissue hypoxia in terrestrial animals. The vertebrate dive response involves vasoconstriction in select tissues, which persists despite profound hypoxia. Using tissues collected from Weddell seals at necropsy, we investigated whether vasoconstriction is aided by downregulation of local hypoxia signaling mechanisms. We focused on NO–soluble guanylyl cyclase (GC)-cGMP signaling, a well-known vasodilatory transduction pathway. Seals have a lower GC protein abundance, activity, and capacity to respond to NO stimulation than do terrestrial mammals. In seal lung homogenates, GC produced less cGMP (20.1 ± 3.7 pmol·mg protein−1·min−1) than the lungs of dogs (−80 ± 144 pmol·mg protein−1·min−1 less than seals), sheep (−472 ± 96), rats (−664 ± 104) or mice (−1,160 ± 104, P < 0.0001). Amino acid sequences of the GC enzyme α-subunits differed between seals and terrestrial mammals, potentially affecting their structure and function. Vasoconstriction in diving Weddell seals is not consistent across tissues; perfusion is maintained in the brain and heart but decreased in other organs such as the kidney. A NO donor increased median GC activity 49.5-fold in the seal brain but only 27.4-fold in the kidney, consistent with the priority of cerebral perfusion during diving. Nos3 expression was high in the seal brain, which could improve NO production and vasodilatory potential. Conversely, Pde5a expression was high in the seal renal artery, which may increase cGMP breakdown and vasoconstriction in the kidney. Taken together, the results of this study suggest that alterations in the NO-cGMP pathway facilitate the diving response.

Download Full-text

(Pro)renin receptor: another member of the system controlled by angiotensin II?

Journal of the Renin-Angiotensin-Aldosterone System ◽

10.1177/1470320311423280 ◽

2011 ◽

Vol 13 (1) ◽

pp. 1-10 ◽

Cited By ~ 5

Author(s):

Luciana G Pereira ◽

Carine P Arnoni ◽

Edgar Maquigussa ◽

Priscila C Cristovam ◽

Juliana Dreyfuss ◽

...

Keyword(s):

Gene Expression ◽

Angiotensin Ii ◽

Mesangial Cells ◽

At1 Receptor ◽

Receptor Internalization ◽

Prorenin Receptor ◽

Receptor Blockers ◽

And Function ◽

At1 Receptor Blockers

The prorenin receptor [(P)RR] is upregulated in the diabetic kidney and has been implicated in the high glucose (HG)-induced overproduction of profibrotic molecules by mesangial cells (MCs), which is mediated by ERK1/2 phosphorylation. The regulation of (P)RR gene transcription and the mechanisms by which HG increases (P)RR gene expression are not fully understood. Because intracellular levels of angiotensin II (AngII) are increased in MCs stimulated with HG, we used this in vitro system to evaluate the possible role of AngII in (P)RR gene expression and function by comparing the effects of AT1 receptor blockers (losartan or candesartan) and (P)RR mRNA silencing (siRNA) in human MCs (HMCs) stimulated with HG. HG induced an increase in (P)RR and fibronectin expression and in ERK1/2 phosphorylation. These effects were completely reversed by (P)RR siRNA and losartan but not by candesartan (an angiotensin receptor blocker that, in contrast to losartan, blocks AT1 receptor internalization). These results suggest that (P)RR gene activity may be controlled by intracellular AngII and that HG-induced ERK1/2 phosphorylation and fibronectin overproduction are primarily induced by (P)RR activation. This relationship between AngII and (P)RR may constitute an additional pathway of MC dysfunction in response to HG stimulation.

Download Full-text

Maturational changes in the regulation of pulmonary vascular tone by nitric oxide in neonatal rats

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00235.2006 ◽

2007 ◽

Vol 293 (5) ◽

pp. L1261-L1270 ◽

Cited By ~ 14

Author(s):

Louis G. Chicoine ◽

Michael L. Paffett ◽

Mark R. Girton ◽

Matthew J. Metropoulus ◽

Mandar S. Joshi ◽

...

Keyword(s):

Nitric Oxide ◽

Guanylyl Cyclase ◽

Vascular Tone ◽

Age Groups ◽

Soluble Guanylyl Cyclase ◽

No Production ◽

Sprague Dawley ◽

No Donor ◽

Early Postnatal Development ◽

Old Rats

Nitric oxide (NO) is an important regulator of vasomotor tone in the pulmonary circulation. We tested the hypothesis that the role NO plays in regulating vascular tone changes during early postnatal development. Isolated, perfused lungs from 7- and 14-day-old Sprague-Dawley rats were studied. Baseline total pulmonary vascular resistance (PVR) was not different between age groups. The addition of KCl to the perfusate caused a concentration-dependent increase in PVR that did not differ between age groups. However, the nitric oxide synthase (NOS) inhibitor Nω-nitro-l-arginine augmented the K+-induced increase in PVR in both groups, and the effect was greater in lungs from 14-day-old rats vs. 7-day-old rats. Lung levels of total endothelial, inducible, and neuronal NOS proteins were not different between groups; however, the production rate of exhaled NO was greater in lungs from 14-day-old rats compared with those of 7-day-old rats. Vasodilation to 0.1 μM of the NO donor spermine NONOate was greater in 14-day lungs than in 7-day lungs, and lung levels of both soluble guanylyl cyclase and cGMP were greater at 14 days than at 7 days. Vasodilation to 100 μM of the cGMP analog 8-(4-chlorophenylthio)guanosine-3′,5′-cyclic monophosphate was greater in 7-day lungs than in 14-day lungs. Our results demonstrate that the pulmonary vascular bed depends more on NO production to modulate vascular tone at 14 days than at 7 days of age. The observed differences in NO sensitivity may be due to maturational increases in soluble guanylyl cyclase protein levels.

Download Full-text

Involvement of guanylyl cyclase and cGMP in the regulation of Mrp2-mediated transport in the proximal tubule

AJP Renal Physiology ◽

10.1152/ajprenal.00443.2003 ◽

2004 ◽

Vol 287 (1) ◽

pp. F33-F38 ◽

Cited By ~ 19

Author(s):

Sylvia Notenboom ◽

David S. Miller ◽

P. Smits ◽

Frans G. M. Russel ◽

Rosalinde Masereeuw

Keyword(s):

Guanylyl Cyclase ◽

Organic Anion ◽

No Donor ◽

Quantitative Image ◽

Cgmp Signaling ◽

Cgmp Analog ◽

A Cell ◽

Protein Isoform ◽

Nos Inhibitor ◽

Oxide Synthase

In killifish renal proximal tubules, endothelin-1 (ET-1), acting through a basolateral ETB receptor, nitric oxide synthase (NOS), and PKC, decreases cell-to-lumen organic anion transport mediated by the multidrug resistance protein isoform 2 (Mrp2). In the present study, we examined the roles of guanylyl cyclase and cGMP in ET signaling to Mrp2. Using confocal microscopy and quantitative image analysis to measure Mrp2-mediated transport of the fluorescent drug fluorescein methotrexate (FL-MTX), we found that oxadiazole quinoxalin (ODQ), an inhibitor of NO-sensitive guanylyl cyclase, blocked ET-1 signaling. ODQ was also effective when signaling was initiated by nephrotoxicants (gentamicin, amikacin, diatrizoate, HgCl2, and CdCl2), which appear to stimulate ET release from the tubules themselves. ODQ blocked the effects of the NO donor sodium nitroprusside but not of the phorbol ester that activates PKC. Exposing tubules to 8-bromo-cGMP (8-BrcGMP), a cell-permeable cGMP analog, decreased luminal FL-MTX accumulation. This effect was abolished by bisindoylmaleimide (BIM), a PKC inhibitor, but not by NG-methyl-l-arginine, a NOS inhibitor. Together, these data indicate that ET regulation of Mrp2 involves activation of guanylyl cyclase and generation of cGMP. Signaling by cGMP follows NO release and precedes PKC activation.

Download Full-text

cGMP: a unique 2nd messenger molecule – recent developments in cGMP research and development

Naunyn-Schmiedeberg s Archives of Pharmacology ◽

10.1007/s00210-019-01779-z ◽

2019 ◽

Vol 393 (2) ◽

pp. 287-302 ◽

Cited By ~ 8

Author(s):

Andreas Friebe ◽

Peter Sandner ◽

Achim Schmidtko

Keyword(s):

Guanylyl Cyclase ◽

Soluble Guanylyl Cyclase ◽

Cell Types ◽

Cyclic Guanosine Monophosphate ◽

Guanosine Monophosphate ◽

Therapeutic Implications ◽

Cellular Effects ◽

Recent Developments ◽

And Function ◽

Messenger Molecule

AbstractCyclic guanosine monophosphate (cGMP) is a unique second messenger molecule formed in different cell types and tissues. cGMP targets a variety of downstream effector molecules and, thus, elicits a very broad variety of cellular effects. Its production is triggered by stimulation of either soluble guanylyl cyclase (sGC) or particulate guanylyl cyclase (pGC); both enzymes exist in different isoforms. cGMP-induced effects are regulated by endogenous receptor ligands such as nitric oxide (NO) and natriuretic peptides (NPs). Depending on the distribution of sGC and pGC and the formation of ligands, this pathway regulates not only the cardiovascular system but also the kidney, lung, liver, and brain function; in addition, the cGMP pathway is involved in the pathogenesis of fibrosis, inflammation, or neurodegeneration and may also play a role in infectious diseases such as malaria. Moreover, new pharmacological approaches are being developed which target sGC- and pGC-dependent pathways for the treatment of various diseases. Therefore, it is of key interest to understand this pathway from scratch, beginning with the molecular basis of cGMP generation, the structure and function of both guanylyl cyclases and cGMP downstream targets; research efforts also focus on the subsequent signaling cascades, their potential crosstalk, and also the translational and, ultimately, the clinical implications of cGMP modulation. This review tries to summarize the contributions to the “9th International cGMP Conference on cGMP Generators, Effectors and Therapeutic Implications” held in Mainz in 2019. Presented data will be discussed and extended also in light of recent landmark findings and ongoing activities in the field of preclinical and clinical cGMP research.

Download Full-text

HNO/cGMP-dependent antihypertrophic actions of isopropylamine-NONOate in neonatal rat cardiomyocytes: potential therapeutic advantages of HNO over NO˙

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00495.2012 ◽

2013 ◽

Vol 305 (3) ◽

pp. H365-H377 ◽

Cited By ~ 24

Author(s):

Jennifer C. Irvine ◽

Nga Cao ◽

Swati Gossain ◽

Amy E. Alexander ◽

Jane E. Love ◽

...

Keyword(s):

Kinase Inhibitor ◽

Oxidant Stress ◽

Pressure Overload ◽

Soluble Guanylyl Cyclase ◽

Neonatal Rat ◽

Cardiomyocyte Hypertrophy ◽

No Donor ◽

Rat Cardiomyocytes ◽

Neonatal Rat Cardiomyocytes ◽

Cgmp Signaling

Nitroxyl (HNO) is a redox congener of NO˙. We now directly compare the antihypertrophic efficacy of HNO and NO˙ donors in neonatal rat cardiomyocytes and compare their contributing mechanisms of actions in this setting. Isopropylamine-NONOate (IPA-NO) elicited concentration-dependent inhibition of endothelin-1 (ET1)-induced increases in cardiomyocyte size, with similar suppression of hypertrophic genes. Antihypertrophic IPA-NO actions were significantly attenuated by l-cysteine (HNO scavenger), Rp-8-pCTP-cGMPS (cGMP-dependent protein kinase inhibitor), and 1-H-(1,2,4)-oxodiazolo-quinxaline-1-one [ODQ; to target soluble guanylyl cyclase (sGC)] but were unaffected by carboxy-PTIO (NO˙ scavenger) or CGRP8–37 (calcitonin gene-related peptide antagonist). Furthermore, IPA-NO significantly increased cardiomyocyte cGMP 3.5-fold (an l-cysteine-sensitive effect) and stimulated sGC activity threefold, without detectable NO˙ release. IPA-NO also suppressed ET1-induced cardiomyocyte superoxide generation. The pure NO˙ donor diethylamine-NONOate (DEA-NO) reproduced these IPA-NO actions but was sensitive to carboxy-PTIO rather than l-cysteine. Although IPA-NO stimulation of purified sGC was preserved under pyrogallol oxidant stress (in direct contrast to DEA-NO), cardiomyocyte sGC activity after either donor was attenuated by this stress. Excitingly IPA-NO also exhibited acute antihypertrophic actions in response to pressure overload in the intact heart. Together these data strongly suggest that IPA-NO protection against cardiomyocyte hypertrophy is independent of both NO˙ and CGRP but rather utilizes novel HNO activation of cGMP signaling. Thus HNO acutely limits hypertrophy independently of NO˙, even under conditions of elevated superoxide. Development of longer-acting HNO donors may thus represent an attractive new strategy for the treatment of cardiac hypertrophy, as stand-alone and/or add-on therapy to standard care.

Download Full-text