scholarly journals A unique choanoflagellate enzyme rhodopsin exhibits light-dependent cyclic nucleotide phosphodiesterase activity

2017 ◽  
Vol 292 (18) ◽  
pp. 7531-7541 ◽  
Author(s):  
Kazuho Yoshida ◽  
Satoshi P. Tsunoda ◽  
Leonid S. Brown ◽  
Hideki Kandori
Keyword(s):  
Cyclic Nucleotides ◽  
Cyclic Nucleotide ◽  
Hydrolytic Activity ◽  
Cyclic Nucleotide Phosphodiesterase ◽  
Hek293 Cells ◽  
Dependent Manner ◽  
Whole Cells ◽  
Microbial Rhodopsin ◽  
Salpingoeca Rosetta ◽  
Photoactivated Adenylyl Cyclase

Photoactivated adenylyl cyclase (PAC) and guanylyl cyclase rhodopsin increase the concentrations of intracellular cyclic nucleotides upon illumination, serving as promising second-generation tools in optogenetics. To broaden the arsenal of such tools, it is desirable to have light-activatable enzymes that can decrease cyclic nucleotide concentrations in cells. Here, we report on an unusual microbial rhodopsin that may be able to meet the demand. It is found in the choanoflagellate Salpingoeca rosetta and contains a C-terminal cyclic nucleotide phosphodiesterase (PDE) domain. We examined the enzymatic activity of the protein (named Rh-PDE) both in HEK293 membranes and whole cells. Although Rh-PDE was constitutively active in the dark, illumination increased its hydrolytic activity 1.4-fold toward cGMP and 1.6-fold toward cAMP, as measured in isolated crude membranes. Purified full-length Rh-PDE displayed maximal light absorption at 492 nm and formed the M intermediate with the deprotonated Schiff base upon illumination. The M state decayed to the parent spectral state in 7 s, producing long-lasting activation of the enzyme domain with increased activity. We discuss a possible mechanism of the Rh-PDE activation by light. Furthermore, Rh-PDE decreased cAMP concentration in HEK293 cells in a light-dependent manner and could do so repeatedly without losing activity. Thus, Rh-PDE may hold promise as a potential optogenetic tool for light control of intracellular cyclic nucleotides (e.g. to study cyclic nucleotide-associated signal transduction cascades).

Cyclic nucleotides, cyclic nucleotide phosphodiesterase, and development in Myxococcus xanthus

1978 ◽  
Vol 24 (12) ◽  
pp. 1475-1481 ◽  
Author(s):  
H. D. McCurdy ◽  
J. Ho ◽  
W. J. Dobson
Keyword(s):  
Fruiting Body ◽  
Cyclic Nucleotides ◽  
Cyclic Nucleotide ◽  
Myxococcus Xanthus ◽  
Dual Role ◽  
Cyclic Nucleotide Phosphodiesterase ◽  
Territory Size ◽  
Subsequent Decrease ◽  
Tentative Model ◽  
Camp Levels

Exogenous cyclic nucleotide phosphodiesterase (PD) accelerated fruiting body (FB) formation and increased territory size of aggregates in Myxococcus xanthus. Both guanosine 3′5′- monophosphate (cGMP) and guanosine 5′-monophosphate (GMP) were antagonistic to the PD effect. Adenosine 3′5′-monophosphate (cAMP) increases FB numbers twofold in the absence but not in the presence of PD. PD induction is not affected by methionine or isoleucine, which inhibit, or by threonine, which stimulates, FB formation. There is an increase and subsequent decrease in cAMP levels during early glycerol-induced microcyst development but 10 mM theophylline or caffeine not only inhibited microcyst development but induced germination in the presence of glycerol.On the basis of these results and the reports of other investigators a tentative model is proposed based on a dual role for cyclic nucleotides in the development in M. xanthus.

scholarly journals Heterogeneity of pulmonary endothelial cyclic nucleotide response to Pseudomonas aeruginosa ExoY infection

2015 ◽  
Vol 309 (10) ◽  
pp. L1199-L1207 ◽  
Author(s):  
K. A. Morrow ◽  
R. Seifert ◽  
V. Kaever ◽  
A. L. Britain ◽  
S. L. Sayner ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Endothelial Cells ◽  
Cyclic Nucleotides ◽  
Cyclic Nucleotide ◽  
Rank Order ◽  
Dependent Manner ◽  
Gap Formation ◽  
Pulmonary Endothelium ◽  
Pulmonary Endothelial Cells ◽  
Intercellular Gaps

Here, we tested the hypothesis that a promiscuous bacterial cyclase synthesizes purine and pyrimidine cyclic nucleotides in the pulmonary endothelium. To test this hypothesis, pulmonary endothelial cells were infected with a strain of the Gram-negative bacterium Pseudomonas aeruginosa that introduces only exoenzyme Y (PA103 ΔexoUexoT::Tc pUCPexoY; ExoY+) via a type III secretion system. Purine and pyrimidine cyclic nucleotides were simultaneously detected using mass spectrometry. Pulmonary artery (PAECs) and pulmonary microvascular (PMVECs) endothelial cells both possess basal levels of four different cyclic nucleotides in the following rank order: cAMP > cUMP ≈ cGMP ≈ cCMP. Endothelial gap formation was induced in a time-dependent manner following ExoY+ intoxication. In PAECs, intercellular gaps formed within 2 h and progressively increased in size up to 6 h, when the experiment was terminated. cGMP concentrations increased within 1 h postinfection, whereas cAMP and cUMP concentrations increased within 3 h, and cCMP concentrations increased within 4 h postinfection. In PMVECs, intercellular gaps did not form until 4 h postinfection. Only cGMP and cUMP concentrations increased at 3 and 6 h postinfection, respectively. PAECs generated higher cyclic nucleotide levels than PMVECs, and the cyclic nucleotide levels increased earlier in response to ExoY+ intoxication. Heterogeneity of the cyclic nucleotide signature in response to P. aeruginosa infection exists between PAECs and PMVECs, suggesting the intracellular milieu in PAECs is more conducive to cNMP generation.

scholarly journals Isoforms of Cyclic Nucleotide Phosphodiesterase PDE3 and Their Contribution to cAMP Hydrolytic Activity in Subcellular Fractions of Human Myocardium

2005 ◽  
Vol 280 (47) ◽  
pp. 39168-39174 ◽  
Author(s):  
Ryan Hambleton ◽  
Judith Krall ◽  
Eliso Tikishvili ◽  
Matthew Honeggar ◽  
Faiyaz Ahmad ◽  
...  
Keyword(s):  
Cyclic Nucleotide ◽  
Hydrolytic Activity ◽  
Human Myocardium ◽  
Subcellular Fractions ◽  
Cyclic Nucleotide Phosphodiesterase

scholarly journals Insulin-Induced Phosphorylation and Activation of Cyclic Nucleotide Phosphodiesterase 3B by the Serine-Threonine Kinase Akt

1999 ◽  
Vol 19 (9) ◽  
pp. 6286-6296 ◽  
Author(s):  
Tadahiro Kitamura ◽  
Yukari Kitamura ◽  
Shoji Kuroda ◽  
Yasuhisa Hino ◽  
Miwa Ando ◽  
...  
Keyword(s):  
Cyclic Nucleotide ◽  
Second Messengers ◽  
Dominant Negative ◽  
Cyclic Nucleotide Phosphodiesterase ◽  
Dominant Negative Mutant ◽  
Dependent Manner ◽  
Serine Threonine Kinase ◽  
Intact Cells ◽  
Threonine Kinase

ABSTRACT Cyclic nucleotide phosphodiesterase (PDE) is an important regulator of the cellular concentrations of the second messengers cyclic AMP (cAMP) and cGMP. Insulin activates the 3B isoform of PDE in adipocytes in a phosphoinositide 3-kinase-dependent manner; however, downstream effectors that mediate signaling to PDE3B remain unknown. Insulin-induced phosphorylation and activation of endogenous or recombinant PDE3B in 3T3-L1 adipocytes have now been shown to be inhibited by a dominant-negative mutant of the serine-threonine kinase Akt, suggesting that Akt is necessary for insulin-induced phosphorylation and activation of PDE3B. Serine-273 of mouse PDE3B is located within a motif (RXRXXS) that is preferentially phosphorylated by Akt. A mutant PDE3B in which serine-273 was replaced by alanine was not phosphorylated either in response to insulin in intact cells or by purified Akt in vitro. In contrast, PDE3B mutants in which alanine was substituted for either serine-296 or serine-421, each of which lies within a sequence (RRXS) preferentially phosphorylated by cAMP-dependent protein kinase, were phosphorylated by Akt in vitro or in response to insulin in intact cells. Moreover, the serine-273 mutant of PDE3B was not activated by insulin when expressed in adipocytes. These results suggest that PDE3B is a physiological substrate of Akt and that Akt-mediated phosphorylation of PDE3B on serine-273 is important for insulin-induced activation of PDE3B.

Chronic Lymphocytic Leukemia Cells Have Increased Expression of Cyclic Nucleotide Phosphodiesterase 7B, Which May Serve as Disease-Specific Therapeutic Target.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 2802-2802
Author(s):  
Lingzhi Zhang ◽  
Fiona Murray ◽  
Joan R. Kanter ◽  
Daisy Chou ◽  
Laura Rassenti ◽  
...  
Keyword(s):  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
Cyclic Nucleotides ◽  
Cyclic Nucleotide ◽  
Lymphocytic Leukemia ◽  
Cyclic Nucleotide Phosphodiesterase ◽  
Target Cells ◽  
Drug Induced ◽  
Induced Apoptosis

Abstract Non-specific inhibition of cyclic nucleotide phosphodiesterase (PDE) in chronic lymphocytic leukemia (CLL) cells leads to intracellular accumulation of cyclic nucleotides, which in turn can sensitize the CLL cells to spontaneous and/or drug-induced apoptosis. Recent studies have identified at least 11 isoforms of PDE, each of which catalyzes the hydrolysis of cAMP and/or cGMP and regulates the intracellular levels of cyclic nucleotides. The finding that various tissues differentially express selected PDE isoforms has prompted development of isoform-specific inhibitors that can selectively increase intracellular levels of cyclic nucleotides in target cells that over-express the inhibited PDE isoform. Accordingly, we examined for differential expression of PDE isoforms by CLL B cells by assessing the levels of mRNA encoding each of the 11 different PDE isoforms in CLL cells and normal blood lymphocytes using quantitative real-time RT-PCR. CLL cell samples (n = 24) and lymphocytes of healthy adults (n = 16) each expressed detectable levels of PDE isoforms 1A, 1B, 2A, 3A, 3B, 4A, 4B, 4C, 4D, 5A, 7A, 7B, 8A, 8B, and 9A. However, we discovered that CLL cells of each patient had significantly higher levels of PDE7B mRNA (2.8-fold to 368-fold) and significantly lower levels of PDE3B mRNA (5-fold to 138 fold) than did lymphocytes from healthy donors (n = 16). As such, the ratios of PDE7B/PDE3B in CLL cell samples were >3 (ranging from 3 to 1019), whereas normal lymphocytes had ratios of < 0.3 (ranging from 0.006 to 0.23). The mean PDE7B/PDE3B ratio for CLL cells (123.6 ± 45.0, S.D., n=24) was significantly higher than that for B-lymphocytes of normal donors (3.8 ± 1.1, n=10) (P<0.0001). Immunoblot analyses demonstrated that CLL cells uniformly expressed high levels of PDE7B and low levels of PDE3B relative to those of normal lymphocytes. Moreover, we found that PDE7B contributed predominantly to the total PDE activity in CLL cells but not in normal lymphocytes. We thus studied effect of a selective PDE7 inhibitor (BRL-50481) on CLL cells and normal lymphocytes in vitro and found that BRL-50481 dose-dependently promoted apoptosis of CLL cells, but not normal lymphocytes. Collectively these findings indicate that CLL B cells selectively over-express PDE7B and under-express PDE3B relative to normal lymphocytes or isolated blood B cells and suggest that selective inhibitors of PDE7B may be effective in the treatment of this disease.

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