scholarly journals KS-505a, an isoform-selective inhibitor of calmodulin-dependent cyclic nucleotide phosphodiesterase

1996 ◽  
Vol 316 (1) ◽  
pp. 311-316 ◽  
Author(s):  
Michio ICHIMURA ◽  
Rika EIKI ◽  
Keiko OSAWA ◽  
Satoshi NAKANISHI ◽  
Hiroshi KASE
Keyword(s):  
Cyclic Nucleotide ◽  
Hippocampal Slices ◽  
Selective Inhibitor ◽  
Binding Domain ◽  
Cyclic Nucleotide Phosphodiesterase ◽  
Intracellular Camp ◽  
Camp Concentration ◽  
Striatal Slices

The effects of KS-505a, a novel microbial metabolite, on the activity of calmodulin-dependent cyclic nucleotide phosphodiesterase (CaM-PDE) were investigated. (1) KS-505a potently inhibited the purified 61 kDa isoenzyme of CaM-PDE from bovine brain and required much higher doses to inhibit the purified 59 kDa isoenzyme of CaM-PDE from bovine heart. The inhibition of both isoenzymes was observed only in the presence of calcium-activated calmodulin (Ca2+/CaM). The IC50 values for the 61 and 59 kDa isoenzymes were 0.17 and 13 μM respectively with 20 μM cAMP as a substrate. (2) Kinetic analysis indicated that the inhibitory mode of KS-505a for the 61 kDa isoenzyme was competitive with respect to Ca2+/CaM; the Ki for KS-505a was 0.089 μM. The inhibition was not competitive with respect to the substrates cAMP or cGMP. (3) KS-505a did not interfere with the interaction between Ca2+/CaM and n-phenyl-1-naphthylamine, a hydrophobic fluorescent probe, nor was it adsorbed to CaM-conjugated gels in the presence of Ca2+, thereby indicating that KS-505a does not bind to Ca2+/CaM. (4) Trypsin-activated 61 kDa isoenzyme, which lacked the Ca2+/CaM-binding domain, was not inhibited by KS-505a at less than micromolar concentrations. Taken together, these results suggest that KS-505a apparently bound to a site in the Ca2+/CaM-binding domain of the 61 kDa isoenzyme and selectively inhibited Ca2+/CaM-activated 61 kDa isoenzyme activity. (5) In rat hippocampal slices, KS-505a at 10 μM increased the intracellular cAMP concentration to approximately three times the basal level, whereas in rat striatal slices it had no effect on the cAMP concentration at concentrations of 1.0–10 μM, suggesting that each CaM-PDE isoenzyme functions differentially in these regions. These results demonstrate that KS-505a is a highly potent selective inhibitor both in vitro and in vivo and distinguishes between subfamily members within the CaM-PDE family.

scholarly journals Cyclic nucleotide phosphodiesterase 1C contributes to abdominal aortic aneurysm

2021 ◽  
Vol 118 (31) ◽  
pp. e2107898118
Author(s):  
Chongyang Zhang ◽  
Hongmei Zhao ◽  
Yujun Cai ◽  
Jian Xiong ◽  
Amy Mohan ◽  
...  
Keyword(s):  
Abdominal Aortic Aneurysm ◽  
Aortic Aneurysm ◽  
Cyclic Nucleotide ◽  
Aortic Aneurysms ◽  
Cyclic Nucleotide Phosphodiesterase ◽  
Causative Role ◽  
Abdominal Aortic

Abdominal aortic aneurysm (AAA) is characterized by aorta dilation due to wall degeneration, which mostly occurs in elderly males. Vascular aging is implicated in degenerative vascular pathologies, including AAA. Cyclic nucleotide phosphodiesterases, by hydrolyzing cyclic nucleotides, play critical roles in regulating vascular structure remodeling and function. Cyclic nucleotide phosphodiesterase 1C (PDE1C) expression is induced in dedifferentiated and aging vascular smooth muscle cells (SMCs), while little is known about the role of PDE1C in aneurysm. We observed that PDE1C was not expressed in normal aorta but highly induced in SMC-like cells in human and murine AAA. In mouse AAA models induced by Angiotensin II or periaortic elastase, PDE1C deficiency significantly decreased AAA incidence, aortic dilation, and elastin degradation, which supported a causative role of PDE1C in AAA development in vivo. Pharmacological inhibition of PDE1C also significantly suppressed preestablished AAA. We showed that PDE1C depletion antagonized SMC senescence in vitro and/or in vivo, as assessed by multiple senescence biomarkers, including senescence-associated β-galactosidase activity, γ-H2AX foci number, and p21 protein level. Interestingly, the role of PDE1C in SMC senescence in vitro and in vivo was dependent on Sirtuin 1 (SIRT1). Mechanistic studies further showed that cAMP derived from PDE1C inhibition stimulated SIRT1 activation, likely through a direct interaction between cAMP and SIRT1, which leads to subsequent up-regulation of SIRT1 expression. Our findings provide evidence that PDE1C elevation links SMC senescence to AAA development in both experimental animal models and human AAA, suggesting therapeutical significance of PDE1C as a potential target against aortic aneurysms.

VEGF-induced HUVEC migration and proliferation are decreased by PDE2 and PDE4 inhibitors

2003 ◽  
Vol 90 (08) ◽  
pp. 334-343 ◽  
Author(s):  
Laure Favot ◽  
Thérèse Keravis ◽  
Vincent Holl ◽  
Alain Bec ◽  
Claire Lugnier
Keyword(s):  
Endothelial Cells ◽  
Cell Cycle Progression ◽  
Umbilical Vein ◽  
Tumor Development ◽  
Chorioallantoic Membrane ◽  
Selective Inhibitor ◽  
Intracellular Camp ◽  
Chick Chorioallantoic Membrane

SummaryMigration and proliferation of endothelial cells in response to VEGF play an important role in angiogenesis associated to pathologies such as atherosclerosis, diabetes and tumor development. Elevation of cAMP in endothelial cells has been shown to inhibit growth factor-induced proliferation. Our hypothesis was that inactivation of cAMP-specific phosphodiesterases (PDEs) would inhibit angiogenesis. The purpose of this study was to evaluate the effect of PDE inhibitors on in vitro and in vivo angiogenesis, using human umbilical vein endothelial cell (HUVEC) and chick chorioallantoic membrane (CAM) models respectively. Here, we report that: 1) PDE2, PDE3, PDE4 and PDE5 are expressed in HUVEC; 2) EHNA (20 µM), PDE2 selective inhibitor, and RP73401 (10 µM), PDE4 selective inhibitor, are able to increase the intracellular cAMP level in HUVEC; 3) EHNA and RP73401 are able to inhibit proliferation, cell cycle progression and migration of HUVEC stimulated by VEGF; 4) these in vitro effects can be mimic by treating HUVEC with the cAMP analogue, 8-Br-cAMP (600 µM); 5) only the association of EHNA and RP73401 inhibits in vivo angiogenesis, indicating that both migration and proliferation must be inhibited. These data strongly suggest that PDE2 and PDE4 represent new potential therapeutic targets in pathological angiogenesis.

5-(2-chloroethyl)-2′-deoxyuridine: A potent and selective inhibitor of herpes viruses in vitro and in vivo

1985 ◽  
Vol 5 ◽  
pp. 21-28 ◽  
Author(s):  
Brigitte Rosenwirth ◽  
Herfried Griengl ◽  
Erich Wanek ◽  
Erik de Clercq
Keyword(s):  
Selective Inhibitor ◽  
Herpes Viruses

scholarly journals Microtubule Actin Cross-Linking Factor (Macf)

1999 ◽  
Vol 147 (6) ◽  
pp. 1275-1286 ◽  
Author(s):  
Conrad L. Leung ◽  
Dongming Sun ◽  
Min Zheng ◽  
David R. Knowles ◽  
Ronald K.H. Liem
Keyword(s):  
Calcium Binding ◽  
Coiled Coil ◽  
Specific Protein ◽  
Binding Domain ◽  
Full Length ◽  
Actin Binding ◽  
Cross Linking ◽  
Actin Binding Domain

We cloned and characterized a full-length cDNA of mouse actin cross-linking family 7 (mACF7) by sequential rapid amplification of cDNA ends–PCR. The completed mACF7 cDNA is 17 kb and codes for a 608-kD protein. The closest relative of mACF7 is the Drosophila protein Kakapo, which shares similar architecture with mACF7. mACF7 contains a putative actin-binding domain and a plakin-like domain that are highly homologous to dystonin (BPAG1-n) at its NH2 terminus. However, unlike dystonin, mACF7 does not contain a coiled–coil rod domain; instead, the rod domain of mACF7 is made up of 23 dystrophin-like spectrin repeats. At its COOH terminus, mACF7 contains two putative EF-hand calcium-binding motifs and a segment homologous to the growth arrest–specific protein, Gas2. In this paper, we demonstrate that the NH2-terminal actin-binding domain of mACF7 is functional both in vivo and in vitro. More importantly, we found that the COOH-terminal domain of mACF7 interacts with and stabilizes microtubules. In transfected cells full-length mACF7 can associate not only with actin but also with microtubules. Hence, we suggest a modified name: MACF (microtubule actin cross-linking factor). The properties of MACF are consistent with the observation that mutations in kakapo cause disorganization of microtubules in epidermal muscle attachment cells and some sensory neurons.

Reconstitution of transcriptional activation domains by reiteration of short peptide segments reveals the modular organization of a glutamine-rich activation domain

1994 ◽  
Vol 14 (9) ◽  
pp. 6056-6067
Author(s):  
M Tanaka ◽  
W Herr
Keyword(s):  
Dna Binding ◽  
Transcriptional Activation ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Activation Domain ◽  
Activation Domains ◽  
Transcriptional Activation Domain ◽  
Pou Domain

The POU domain activator Oct-2 contains an N-terminal glutamine-rich transcriptional activation domain. An 18-amino-acid segment (Q18III) from this region reconstituted a fully functional activation domain when tandemly reiterated and fused to either the Oct-2 or GAL4 DNA-binding domain. A minimal transcriptional activation domain likely requires three tandem Q18III segments, because one or two tandem Q18III segments displayed little activity, whereas three to five tandem segments were active and displayed increasing activity with increasing copy number. As with natural Oct-2 activation domains, in our assay a reiterated activation domain required a second homologous or heterologous activation domain to stimulate transcription effectively when fused to the Oct-2 POU domain. These results suggest that there are different levels of synergy within and among activation domains. Analysis of reiterated activation domains containing mutated Q18III segments revealed that leucines and glutamines, but not serines or threonines, are critical for activity in vivo. Curiously, several reiterated activation domains that were inactive in vivo were active in vitro, suggesting that there are significant functional differences in our in vivo and in vitro assays. Reiteration of a second 18-amino-acid segment from the Oct-2 glutamine-rich activation domain (Q18II) was also active, but its activity was DNA-binding domain specific, because it was active when fused to the GAL4 than to the Oct-2 DNA-binding domain. The ability of separate short peptide segments derived from a single transcriptional activation domain to activate transcription after tandem reiteration emphasizes the flexible and modular nature of a transcriptional activation domain.

NUANCE, a giant protein connecting the nucleus and actin cytoskeleton

2002 ◽  
Vol 115 (15) ◽  
pp. 3207-3222 ◽  
Author(s):  
Yen-Yi Zhen ◽  
Thorsten Libotte ◽  
Martina Munck ◽  
Angelika A. Noegel ◽  
Elena Korenbaum
Keyword(s):  
Actin Cytoskeleton ◽  
Transmembrane Domain ◽  
Coiled Coil ◽  
Peripheral Blood Lymphocytes ◽  
Binding Domain ◽  
Actin Binding ◽  
Actin Binding Domain ◽  
Microfilament System

NUANCE (NUcleus and ActiN Connecting Element) was identified as a novel protein with an α-actinin-like actin-binding domain. A human 21.8 kb cDNA of NUANCE spreads over 373 kb on chromosome 14q22.1-q22.3. The cDNA sequence predicts a 796 kDa protein with an N-terminal actin-binding domain, a central coiled-coil rod domain and a predicted C-terminal transmembrane domain. High levels of NUANCE mRNA were detected in the kidney, liver,stomach, placenta, spleen, lymphatic nodes and peripheral blood lymphocytes. At the subcellular level NUANCE is present predominantly at the outer nuclear membrane and in the nucleoplasm. Domain analysis shows that the actin-binding domain binds to Factin in vitro and colocalizes with the actin cytoskeleton in vivo as a GFP-fusion protein. The C-terminal transmembrane domain is responsible for the targeting the nuclear envelope. Thus, NUANCE is the firstα-actinin-related protein that has the potential to link the microfilament system with the nucleus.

Chloroquine stimulates absorption and inhibits secretion of ileal water and electrolytes

1982 ◽  
Vol 243 (2) ◽  
pp. G117-G126
Author(s):  
R. Fogel ◽  
G. W. Sharp ◽  
M. Donowitz
Keyword(s):  
Cholera Toxin ◽  
Calcium Content ◽  
Intracellular Camp ◽  
Rabbit Ileum ◽  
Camp Content ◽  
Serosal Surface ◽  
Ileal Absorption ◽  
Camp Levels

The effects of chloroquine diphosphate, a drug with "'membrane-stabilizing" properties, were studied on basal ileal absorption and on ileal secretion induced by increased intracellular cAMP levels and calcium (serotonin). The studies were performed on rat (in vivo) and rabbit ileum (in vitro). Intraluminal chloroquine (10(-4) M) reversed cholera toxin- and theophylline-induced secretion in rat ileum but did not alter the cholera toxin- and theophylline-induced increases in cAMP content. Addition of chloroquine (10(-4) M) to the mucosal surface of rabbit ileum did not alter basal active electrolyte transport or the serotonin-induced decreased Na and Cl absorption but inhibited the theophylline-induced C1 secretion. Addition of chloroquine (10(-4)) M) to the serosal surface stimulated net Na and Cl absorption. This effect may involve intracellular calcium. Chloroquine increased the rabbit ileal calcium content and decreased 45Ca2+ influx from the serosal surface. Both the mucosal and serosal effects of chloroquine described led to a net increase in absorptive function of the intestine and should prove useful in developing treatment of diarrheal diseases.

scholarly journals Bidirectional in vivo structural dendritic spine plasticity revealed by two-photon glutamate uncaging in the mouse neocortex

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Jun Noguchi ◽  
Akira Nagaoka ◽  
Tatsuya Hayama ◽  
Hasan Ucar ◽  
Sho Yagishita ◽  
...  
Keyword(s):  
Dendritic Spines ◽  
Time Course ◽  
Hippocampal Slices ◽  
Low Frequency ◽  
Structural Plasticity ◽  
Experimental Conditions ◽  
Two Photon ◽  
Spine Plasticity

Abstract Most excitatory synapses in the brain form on dendritic spines. Two-photon uncaging of glutamate is widely utilized to characterize the structural plasticity of dendritic spines in brain slice preparations in vitro. In the present study, glutamate uncaging was used to investigate spine plasticity, for the first time, in vivo. A caged glutamate compound was applied to the surface of the mouse visual cortex in vivo, revealing the successful induction of spine enlargement by repetitive two-photon uncaging in a magnesium free solution. Notably, this induction occurred in a smaller fraction of spines in the neocortex in vivo (22%) than in hippocampal slices (95%). Once induced, the time course and mean long-term enlargement amplitudes were similar to those found in hippocampal slices. However, low-frequency (1–2 Hz) glutamate uncaging in the presence of magnesium caused spine shrinkage in a similar fraction (35%) of spines as in hippocampal slices, though spread to neighboring spines occurred less frequently than it did in hippocampal slices. Thus, the structural plasticity may occur similarly in the neocortex in vivo as in hippocampal slices, although it happened less frequently in our experimental conditions.

Sign in / Sign up

Export Citation Format

Share Document