scholarly journals Functional Analysis of the Expanded Phosphodiesterase Gene Family in Toxoplasma gondii Tachyzoites

2021 ◽  
Author(s):  
William J Moss ◽  
Caitlyn E Patterson ◽  
Alexander K Jochmans ◽  
Kevin M Brown
Keyword(s):  
Cyclic Nucleotide ◽  
Biochemical Analysis ◽  
Life Stage ◽  
Master Regulators ◽  
Growth Defects ◽  
Expression Studies ◽  
Cgmp Signaling ◽  
And Function ◽  
Camp Levels ◽  
Severe Growth

Toxoplasma motility is both activated and suppressed by 3’-5’ cyclic nucleotide signaling. Cyclic GMP (cGMP) signaling through TgPKG activates motility, whereas cyclic AMP (cAMP) signaling through TgPKAc1 inhibits motility. Despite being master regulators of motility, it is unclear how cGMP and cAMP levels are maintained in Toxoplasma. Phosphodiesterases (PDEs) are known to inactivate cyclic nucleotides and are highly expanded in the Toxoplasma genome. Here we utilized an auxin-inducible degron system to analyze the expression and function of the 18-member TgPDE family in tachyzoites, the virulent life stage of Toxoplasma. We detected the expression of 11 of 18 TgPDEs by immunofluorescence microscopy or immunoblotting, confirming prior expression studies. We performed a knockdown screen of the TgPDE family and identified four TgPDEs that contribute to lytic Toxoplasma growth (TgPDE1, TgPDE2, TgPDE5, and TgPDE9). Loss of TgPDE1 and TgPDE2 caused severe growth defects, prompting further investigation. TgPDE1 displayed a plasma membrane/cytomembranous distribution, whereas TgPDE2 displayed an endoplasmic reticulum-like distribution. Biochemical analysis of TgPDE1 and TgPDE2 purified from Toxoplasma lysates revealed that they are active phosphodiesterases. TgPDE1 was capable of hydrolyzing both cGMP and cAMP, whereas TgPDE2 was cAMP-specific. Interactome studies of TgPDE1 and TgPDE2 indicated that they do not physically interact with each other or other TgPDEs but may be regulated by kinases and proteases. Our studies have identified TgPDE1 and TgPDE2 as central regulators of tachyzoite cyclic nucleotide levels and enable future studies aimed at determining how these enzymes are regulated and cooperate to control Toxoplasma motility.

Ticlopidine and Clopidogrel (SR 25990C) Selectively Neutralize ADP Inhibition of PGE1-Activated Platelet Adenylate Cyclase in Rats and Rabbits

1991 ◽  
Vol 65 (02) ◽  
pp. 186-190 ◽  
Author(s):  
G Defreyn ◽  
C Gachet ◽  
P Savi ◽  
F Driot ◽  
J P Cazenave ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Adenylate Cyclase ◽  
Direct Effect ◽  
Cyclic Nucleotide ◽  
Camp Accumulation ◽  
Rabbit Platelets ◽  
Camp Levels ◽  
Kinetics Of ◽  
Camp Elevation ◽  
Rat Platelets

SummaryTiclopidine and its potent analogue, clopidogrel, are powerful inhibitors of ADP-induced platelet aggregation. In order to improve the understanding of this ADP-selectivity, we studied the effect of these compounds on PGE1-stimulated adenylate cyclase and on the inhibition of this enzyme by ADP, epinephrine and thrombin. Neither drug changed the basal cAMP levels nor the kinetics of cAMP accumulation upon PGEj-stimulation in rat or rabbit platelets, which excludes any direct effect on adenylate cyclase or on cyclic nucleotide phosphodiesterase. However, the drop in cAMP levels observed after addition of ADP to PGEr stimulated control platelets was inhibited in platelets from treated animals. In contrast, the drop in cAMP levels produced by epinephrine was not prevented by either drug in rabbit platelets. In rat platelets, thrombin inhibited the PGEX-induced cAMP elevation but this effect seems to be entirely mediated by the released ADP. Under these conditions, it was not surprising to find that clopidogrel also potently inhibited that effect of thrombin on platelet adenylate cyclase. In conclusion, ticlopidine and clopidogrel selectively neutralize the ADP inhibition of PGEr activated platelet adenylate cyclase in rats and rabbits.

scholarly journals Vitamin D metabolism, sex hormones, and male reproductive function

Reproduction ◽  
2012 ◽  
Vol 144 (2) ◽  
pp. 135-152 ◽  
Author(s):  
Martin Blomberg Jensen
Keyword(s):  
Vitamin D ◽  
Association Studies ◽  
Reproductive Function ◽  
Hormone Production ◽  
Vitamin D Metabolism ◽  
Mediated Effects ◽  
Male Reproductive Function ◽  
Expression Studies ◽  
Metabolizing Enzyme ◽  
And Function

The spectrum of vitamin D (VD)-mediated effects has expanded in recent years, and VD is now recognized as a versatile signaling molecule rather than being solely a regulator of bone health and calcium homeostasis. One of the recently identified target areas of VD is male reproductive function. The VD receptor (VDR) and the VD metabolizing enzyme expression studies documented the presence of this system in the testes, mature spermatozoa, and ejaculatory tract, suggesting that both systemic and local VD metabolism may influence male reproductive function. However, it is still debated which cell is the main VD target in the testis and to what extent VD is important for sex hormone production and function of spermatozoa. This review summarizes descriptive studies on testicular VD metabolism and spatial distribution of VDR and the VD metabolizing enzymes in the mammalian testes and discusses mechanistic and association studies conducted in animals and humans. The reviewed evidence suggests some effects of VD on estrogen and testosterone biosynthesis and implicates involvement of both systemic and local VD metabolism in the regulation of male fertility potential.

Mechanisms of cyclic nucleotide-induced relaxation in canine tracheal smooth muscle

1995 ◽  
Vol 268 (3) ◽  
pp. L407-L413 ◽  
Author(s):  
I. McGrogan ◽  
S. Lu ◽  
S. Hipworth ◽  
L. Sormaz ◽  
R. Eng ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Cyclic Nucleotide ◽  
Cyclopiazonic Acid ◽  
Inhibitory Effect ◽  
Beta Agonist ◽  
Cyclic Monophosphate ◽  
Median Effective Concentration ◽  
Camp Levels ◽  
Ca2 Channels

The effects of exogeneous cyclopiazonic acid (CPA, 10 microM), a selective inhibitor of the sarcoplasmic reticulum (SR) Ca2+ adenosinetriphosphatase, on cyclic nucleotide-induced relaxations of canine airway smooth muscle were examined. Strips of tracheal muscle were precontracted with carbachol (50% median effective concentration, 0.1 microM) or with 60 mM KCl. The beta-agonist isoproterenol (ISO, 10 microM) relaxed the tissue by approximately 50%. The relaxation was reduced in the presence of CPA when L-type Ca2+ channels were available but not when these were blocked by 0.1 microM nifedipine. Forskolin (1.0 microM), an adenylate cyclase activator, was less effective at inhibiting the contraction than ISO, and addition of CPA did not block its inhibitory effect as effectively as when ISO was used. Radioimmunoassay indicated that both these agents raised adenosine 3',5'-cyclic monophosphate (cAMP) levels to the same degree. Very little relaxation of the precontracted smooth muscle was elicited by 3 mM 8-bromo-adenosine 3',5'-cyclic monophosphate (8-BrcAMP), and addition of CPA had no effect. Sodium nitroprusside (100 microM) and 8-bromo-guanosine 3',5'-cyclic monophosphate (10 mM) inhibited contraction to a greater degree than any agent that raised cAMP. These inhibitions were greatly reduced in the presence of CPA when L-type Ca2+ channels were available. We conclude that pumping of Ca2+ into SR plays a major role guanosine 3',5'-cyclic monophosphate-produced but not cAMP-induced relaxation; L-type Ca2+ channels must be available for the relaxant role of Ca2+ pumping into the SR to be expressed; and ISO-induced relaxation may not involve primarily elevation of the cAMP.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals The encapsulin from Thermatoga maritima is a flavoprotein with a symmetry matched ferritin-like cargo protein

2021 ◽  
Author(s):  
Benjamin J LaFrance ◽  
Caleb Cassidy-Amstutz ◽  
Robert J Nichols ◽  
Luke M Oltrogge ◽  
Eva Nogales ◽  
...  
Keyword(s):  
Enzymatic Activity ◽  
Biochemical Analysis ◽  
Active Role ◽  
Structure And Function ◽  
Redox Active ◽  
Cargo Protein ◽  
Cargo Proteins ◽  
And Function ◽  
Flavin Binding ◽  
Unique Chemical

Bacterial nanocompartments, also known as encapsulins, are an emerging class of protein-based "organelles" found in bacteria and archaea. Encapsulins are virus-like icosahedral particles comprising a ~25-50 nm shell surrounding a specific cargo enzyme. Compartmentalization is thought to create a unique chemical environment to facilitate catalysis and isolate toxic intermediates. Many questions regarding nanocompartment structure-function remain unanswered, including how shell symmetry dictates cargo loading and to what extent the shell facilitates enzymatic activity. Here, we explore these questions using the model T. maritima nanocompartment known to encapsulate a redox-active ferritin-like protein. Biochemical analysis revealed the encapsulin shell to possess a flavin binding site located at the interface between capsomere subunits, suggesting the shell may play a direct and active role in the function of the encapsulated cargo. Furthermore, we used cryoEM to show that cargo proteins use a form of symmetry-matching to facilitate encapsulation and define stoichiometry. In the case of the T. maritima encapsulin, the decameric cargo protein with 5-fold symmetry preferentially binds to the pentameric-axis of the icosahedral shell. Taken together, these observations suggest the shell is not simply a passive barrier-it also plays a significant role in the structure and function of the cargo enzyme.

scholarly journals Plasma cyclic nucleotide levels in acute leukemia patients

Blood ◽  
1987 ◽  
Vol 69 (6) ◽  
pp. 1613-1616 ◽  
Author(s):  
M Peracchi ◽  
V Toschi ◽  
F Bamonti-Catena ◽  
L Lombardi ◽  
B Bareggi ◽  
...  
Keyword(s):  
Complete Remission ◽  
Cyclic Amp ◽  
Acute Leukemia ◽  
Cyclic Nucleotide ◽  
Normal Subjects ◽  
Response To Treatment ◽  
Active Stage ◽  
Normal Range ◽  
Plasma Camp ◽  
Camp Levels

Abstract To verify the clinical usefulness of extracellular cyclic nucleotide determination as a tumor marker, plasma cyclic AMP (cAMP) and cyclic GMP (cGMP) levels were measured in 70 normal subjects and 173 acute leukemia patients studied in different stages of their disease. Mean plasma cAMP levels were similar in leukemic and normal subjects, although in 48 patients in the active stage of the disease, first diagnosis, or relapse, the cAMP values were below the normal range, and most of these patients failed to respond to chemotherapy. Plasma cGMP levels were markedly elevated in untreated patients, normalized in all patients who attained complete remission, and increased promptly to pretreatment values in patients who relapsed, suggesting that their determination may be useful to monitor the patients' response to treatment.

Arabidopsis mtHSC70-1 plays important roles in the establishment of COX-dependent respiration and redox homeostasis

2019 ◽  
Vol 70 (20) ◽  
pp. 5575-5590 ◽  
Author(s):  
Shan-Shan Wei ◽  
Wei-Tao Niu ◽  
Xiao-Ting Zhai ◽  
Wei-Qian Liang ◽  
Meng Xu ◽  
...  
Keyword(s):  
Alternative Oxidase ◽  
Redox Homeostasis ◽  
Superoxide Dismutase 1 ◽  
Respiratory Pathway ◽  
Cellular Processes ◽  
Growth Defects ◽  
Alternative Respiratory Pathway ◽  
Abnormal Mitochondria ◽  
Shock Proteins ◽  
Severe Growth

Abstract The 70 kDa heat shock proteins function as molecular chaperones and are involved in diverse cellular processes. However, the functions of the plant mitochondrial HSP70s (mtHSC70s) remain unclear. Severe growth defects were observed in the Arabidopsis thaliana mtHSC70-1 knockout lines, mthsc70-1a and mthsc70-1b. Conversely, the introduction of the mtHSC70-1 gene into the mthsc70-1a background fully reversed the phenotypes, indicating that mtHSC70-1 is essential for plant growth. The loss of mtHSC70-1 functions resulted in abnormal mitochondria and alterations to respiration because of an inhibition of the cytochrome c oxidase (COX) pathway and the activation of the alternative respiratory pathway. Defects in COX assembly were observed in the mtHSC70-1 knockout lines, leading to decreased COX activity. The mtHSC70-1 knockout plants have increased levels of reactive oxygen species (ROS). The introduction of the Mn-superoxide dismutase 1 (MSD1) or the catalase 1 (CAT1) gene into the mthsc70-1a plants decreased ROS levels, reduced the expression of alternative oxidase, and partially rescued growth. Taken together, our data suggest that mtHSC70-1 plays important roles in the establishment of COX-dependent respiration.

The cGMP system: components and function

2020 ◽  
Vol 401 (4) ◽  
pp. 447-469 ◽  
Author(s):  
Franz Hofmann
Keyword(s):  
Cyclic Guanosine Monophosphate ◽  
Guanosine Monophosphate ◽  
Signaling System ◽  
Pde Inhibitors ◽  
Mammalian Tissues ◽  
System Components ◽  
Cgmp Signaling ◽  
Dependent Protein ◽  
Successful Approach ◽  
And Function

AbstractThe cyclic guanosine monophosphate (cGMP) signaling system is one of the most prominent regulators of a variety of physiological and pathophysiological processes in many mammalian and non-mammalian tissues. Targeting this pathway by increasing cGMP levels has been a very successful approach in pharmacology as shown for nitrates, phosphodiesterase (PDE) inhibitors and stimulators of nitric oxide-guanylyl cyclase (NO-GC) and particulate GC (pGC). This is an introductory review to the cGMP signaling system intended to introduce those readers to this system, who do not work in this area. This article does not intend an in-depth review of this system. Signal transduction by cGMP is controlled by the generating enzymes GCs, the degrading enzymes PDEs and the cGMP-regulated enzymes cyclic nucleotide-gated ion channels, cGMP-dependent protein kinases and cGMP-regulated PDEs. Part A gives a very concise introduction to the components. Part B gives a very concise introduction to the functions modulated by cGMP. The article cites many recent reviews for those who want a deeper insight.

scholarly journals Redundant Roles of Tead1 and Tead2 in Notochord Development and the Regulation of Cell Proliferation and Survival

2008 ◽  
Vol 28 (10) ◽  
pp. 3177-3189 ◽  
Author(s):  
Atsushi Sawada ◽  
Hiroshi Kiyonari ◽  
Kanako Ukita ◽  
Noriyuki Nishioka ◽  
Yu Imuta ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Paraxial Mesoderm ◽  
Mouse Development ◽  
Lateral Plate ◽  
Growth Defects ◽  
Mouse Mutants ◽  
Severe Growth

ABSTRACT Four members of the TEAD/TEF family of transcription factors are expressed widely in mouse embryos and adult tissues. Although in vitro studies have suggested various roles for TEAD proteins, their in vivo functions remain poorly understood. Here we examined the role of Tead genes by generating mouse mutants for Tead1 and Tead2. Tead2 −/− mice appeared normal, but Tead1 −/−; Tead2 −/− embryos died at embryonic day 9.5 (E9.5) with severe growth defects and morphological abnormalities. At E8.5, Tead1 −/−; Tead2 −/− embryos were already small and lacked characteristic structures such as a closed neural tube, a notochord, and somites. Despite these overt abnormalities, differentiation and patterning of the neural plate and endoderm were relatively normal. In contrast, the paraxial mesoderm and lateral plate mesoderm were displaced laterally, and a differentiated notochord was not maintained. These abnormalities and defects in yolk sac vasculature organization resemble those of mutants for Yap, which encodes a coactivator of TEAD proteins. Moreover, we demonstrated genetic interactions between Tead1 and Tead2 and Yap. Finally, Tead1 −/−; Tead2 −/− embryos showed reduced cell proliferation and increased apoptosis. These results suggest that Tead1 and Tead2 are functionally redundant, use YAP as a major coactivator, and support notochord maintenance as well as cell proliferation and survival in mouse development.

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