Overexpression of a directed mutant of 14-3-3ω in Arabidopsis leaves affects phosphorylation and protein content of nitrate reductaseThis paper is one of a selection published in a Special Issue comprising papers presented at the 50th Annual Meeting of the Canadian Society of Plant Physiologists (CSPP) held at the University of Ottawa, Ontario, in June 2008.

Botany ◽  
2009 ◽  
Vol 87 (7) ◽  
pp. 691-701 ◽  
Author(s):  
Man-Ho Oh ◽  
Joan L. Huber ◽  
Wei Shen ◽  
Gurdeep S. Athwal ◽  
Xia Wu ◽  
...  
Keyword(s):  
Defense Responses ◽  
Cellular Proteins ◽  
Signaling Proteins ◽  
Glutathione S Transferase ◽  
Independent Manner ◽  
Client Proteins ◽  
The University

The 14-3-3 family of proteins are highly conserved signaling proteins in eukaryotes that bind to their client proteins, usually through specific phosphorylated target sequences. While the 14-3-3 proteins are thought to interact with a wide array of cellular proteins, there have been few studies addressing the in-vivo role of 14-3-3. As one approach to study this in-vivo role, we generated transgenic Arabidopsis plants constitutively overexpressing a directed mutant of 14-3-3 isoform ω that inhibits phosphorylated nitrate reductase (pNR) in a largely divalent-cation-independent manner in vitro. The transgenic plants had increased relative phosphorylation of NR at the regulatory Ser-534 site and decreased NR activity measured in the presence of 5 mmol·L–1 MgCl2 relative to nontransgenic plants. In addition, total NR protein was increased and the protein half-life was increased about two-fold. Two-dimensional difference gel electrophoresis analysis of proteins extracted from leaves of plants expressing the mutant 14-3-3 identified numerous cellular proteins that were altered in abundance. In particular, several β-glucosidase and glutathione S-transferase isoforms were decreased in abundance relative to wild type plants suggesting a possible alteration in stress or defense responses.

scholarly journals A Possible Role of Allatostatin C in Inhibiting Ecdysone Biosynthesis Revealed in the Mud Crab Scylla paramamosain

2021 ◽  
Vol 8 ◽  
Author(s):  
An Liu ◽  
Wenyuan Shi ◽  
Dongdong Lin ◽  
Haihui Ye
Keyword(s):  
Scylla Paramamosain ◽  
Dependent Manner ◽  
Mud Crab ◽  
Methyl Farnesoate ◽  
Independent Manner ◽  
Wide Range ◽  
Negative Effect

C-type allatostatins (C-type ASTs) are a family of structurally related neuropeptides found in a wide range of insects and crustaceans. To date, the C-type allatostatin receptor in crustaceans has not been deorphaned, and little is known about its physiological functions. In this study, we aimed to functionally define a C-type ASTs receptor in the mud crab, Scylla paramamosian. We showed that C-type ASTs receptor can be activated by ScypaAST-C peptide in a dose-independent manner and by ScypaAST-CCC peptide in a dose-dependent manner with an IC50 value of 6.683 nM. Subsequently, in vivo and in vitro experiments were performed to investigate the potential roles of ScypaAST-C and ScypaAST-CCC peptides in the regulation of ecdysone (20E) and methyl farnesoate (MF) biosynthesis. The results indicated that ScypaAST-C inhibited biosynthesis of 20E in the Y-organ, whereas ScypaAST-CCC had no effect on the production of 20E. In addition, qRT-PCR showed that both ScypaAST-C and ScypaAST-CCC significantly decreased the level of expression of the MF biosynthetic enzyme gene in the mandibular organ, suggesting that the two neuropeptides have a negative effect on the MF biosynthesis in mandibular organs. In conclusion, this study provided new insight into the physiological roles of AST-C in inhibiting ecdysone biosynthesis. Furthermore, it was revealed that AST-C family peptides might inhibit MF biosynthesis in crustaceans.

Abstract 140: Anti-inflammatory and Anti-atherogenic Role of Bmp Receptor II In Endothelial Cells

2013 ◽  
Vol 33 (suppl_1) ◽  
Author(s):  
Chanwoo Kim ◽  
Hannah Song ◽  
Sandeep Kumar ◽  
Douglas Nam ◽  
Hyuk Sang Kwon ◽  
...  
Keyword(s):  
Endothelial Dysfunction ◽  
Intracellular Signaling ◽  
Independent Manner ◽  
Disturbed Flow ◽  
Bmp Receptors ◽  
Endothelial Inflammation ◽  
Anti Inflammatory

Atherosclerosis is a multifactorial disease that arises from a combination of endothelial dysfunction and inflammation, occurring preferentially in arterial regions exposed to disturbed flow. Bone morphogenic protein-4 (BMP4) produced by disturbed flow induces inflammation, endothelial dysfunction and hypertension, suggesting the importance of BMPs in vascular biology and disease. BMPs bind to two different types of BMP receptors (BMPRI and II) to instigate intracellular signaling. Increasing evidences suggest a correlative role of BMP4 and atherosclerosis, but the role of BMP receptors especially BMPRII in atherosclerosis is still unclear and whether knockdown of BMPRII is the cause or the consequence of atherosclerosis is still not known. It is therefore, imperative to investigate the mechanisms by which BMPRII expression is modulated and its ramifications in atherosclerosis. Initially, we expected that knockdown of BMPRII will result in loss of pro-atherogenic BMP4 signaling and will thereby prevent atherosclerosis. Contrarily, we found that loss of BMPRII expression causes endothelial inflammation and atherosclerosis. Using BMPRII siRNA and BMPRII +/- mice, we found that BMPRII knockdown induces endothelial inflammation in a BMP-independent manner via mechanisms involving reactive oxygen species (ROS), NFκB, and NADPH oxidases. Further, BMPRII +/- ApoE -/- mice develop accelerated atherosclerosis compared to BMPRII +/+ ApoE -/- mice, suggesting loss of BMPRII may induce atherosclerosis. Interestingly, we found that multiple pro-atherogenic stimuli such as hypercholesterolemia, disturbed flow, pro-hypertensive angiotensin II, and pro-inflammatory cytokine, TNFα, downregulate BMPRII expression in endothelium, while anti-atherogenic stimuli such as stable flow and statin treatment upregulate its expression, both in vivo and in vitro . Moreover, we found that BMPRII expression is significantly diminished in human coronary advanced atherosclerotic lesions. These results suggest that BMPRII is a critical, anti-inflammatory and anti-atherogenic protein that is commonly targeted by multiple pro- and anti-atherogenic factors. BMPRII could be used as a novel diagnostic and therapeutic target in atherosclerosis.

scholarly journals cGAS-mediated autophagy protects the liver from ischemia-reperfusion injury independently of STING

2018 ◽  
Vol 314 (6) ◽  
pp. G655-G667 ◽  
Author(s):  
Zhao Lei ◽  
Meihong Deng ◽  
Zhongjie Yi ◽  
Qian Sun ◽  
Richard A. Shapiro ◽  
...  
Keyword(s):  
Ischemia Reperfusion Injury ◽  
Ischemia Reperfusion ◽  
Adenosine Monophosphate ◽  
Cyclic Guanosine Monophosphate ◽  
Protective Role ◽  
Guanosine Monophosphate ◽  
Independent Manner

Liver ischemia-reperfusion (I/R) injury occurs through induction of oxidative stress and release of damage-associated molecular patterns (DAMPs), including cytosolic DNA released from dysfunctional mitochondria or from the nucleus. Cyclic guanosine monophosphate–adenosine monophosphate (cGAMP) synthase (cGAS) is a cytosolic DNA sensor known to trigger stimulator of interferon genes (STING) and downstream type 1 interferon (IFN-I) pathways, which are pivotal innate immune system responses to pathogen. However, little is known about the role of cGAS/STING in liver I/R injury. We subjected C57BL/6 (WT), cGAS knockout (cGAS−/−), and STING-deficient (STINGgt/gt) mice to warm liver I/R injury and that found cGAS−/− mice had significantly increased liver injury compared with WT or STINGgt/gt mice, suggesting a protective effect of cGAS independent of STING. Liver I/R upregulated cGAS in vivo and also in vitro in hepatocytes subjected to anoxia/reoxygenation (A/R). We confirmed a previously published finding that hepatocytes do not express STING under normoxic conditions or after A/R. Hepatocytes and liver from cGAS−/− mice had increased cell death and reduced induction of autophagy under hypoxic conditions as well as increased apoptosis. Protection could be restored in cGAS−/− hepatocytes by overexpression of cGAS or by pretreatment of mice with autophagy inducer rapamycin. Our findings indicate a novel protective role for cGAS in the regulation of autophagy during liver I/R injury that occurs independently of STING. NEW & NOTEWORTHY Our studies are the first to document the important role of cGAS in the acute setting of sterile injury induced by I/R. Specifically, we provide evidence that cGAS protects liver from I/R injury in a STING-independent manner.

In vitro and in vivo effects of progesterone on Trichinella spiralis newborn larvae

Parasitology ◽  
2005 ◽  
Vol 131 (2) ◽  
pp. 255-259 ◽  
Author(s):  
G. G. NUÑEZ ◽  
T. GENTILE ◽  
S. N. COSTANTINO ◽  
M. I. SARCHI ◽  
S. M. VENTURIELLO
Keyword(s):  
Trichinella Spiralis ◽  
Parasite Load ◽  
Ovariectomized Rats ◽  
Independent Manner ◽  
Pregnant Rats ◽  
Peritoneal Cells ◽  
In Vivo Effects

We have previously demonstrated that during pregnancy there exists an increased parasiticide activity against Trichinella spiralis newborn larvae (NBL) in infected rats. In this work we analysed the contribution of peritoneal cells from non-infected pregnant rats to the mortality of the NBL in cytotoxicity assays, and evaluated the role of progesterone in this effector mechanism. Our findings suggest that progesterone can induce activation of effector peritoneal cells to destroy the NBL in a rapid and antibody-independent manner. The administration of progesterone to ovariectomized rats also led to a significant decrease in the parasite load of the animals, thus demonstrating that progesterone induces the increase of the parasiticide activity of the leukocytes involved in the mechanisms of NBL death.

scholarly journals Definition of the minimal fragments of Sti1 required for dimerization, interaction with Hsp70 and Hsp90 and in vivo functions

2007 ◽  
Vol 404 (1) ◽  
pp. 159-167 ◽  
Author(s):  
Gary Flom ◽  
Robert H. Behal ◽  
Luke Rosen ◽  
Douglas G. Cole ◽  
Jill L. Johnson
Keyword(s):  
Current Model ◽  
Heat Shock Protein 90 ◽  
C Terminus ◽  
Client Proteins ◽  
Definition Of ◽  
Necessary And Sufficient ◽  
In Vitro Interaction

The molecular chaperone Hsp (heat-shock protein) 90 is critical for the activity of diverse cellular client proteins. In a current model, client proteins are transferred from Hsp70 to Hsp90 in a process mediated by the co-chaperone Sti1/Hop, which may simultaneously interact with Hsp70 and Hsp90 via separate TPR (tetratricopeptide repeat) domains, but the mechanism and in vivo importance of this function is unclear. In the present study, we used truncated forms of Sti1 to determine the minimal regions required for the Hsp70 and Hsp90 interaction, as well as Sti1 dimerization. We found that both TPR1 and TPR2B contribute to the Hsp70 interaction in vivo and that mutations in both TPR1 and TPR2B were required to disrupt the in vitro interaction of Sti1 with the C-terminus of the Hsp70 Ssa1. The TPR2A domain was required for the Hsp90 interaction in vivo, but the isolated TPR2A domain was not sufficient for the Hsp90 interaction unless combined with the TPR2B domain. However, isolated TPR2A was both necessary and sufficient for purified Sti1 to migrate as a dimer in solution. The DP2 domain, which is essential for in vivo function, was dispensable for the Hsp70 and Hsp90 interaction, as well as Sti1 dimerization. As evidence for the role of Sti1 in mediating the interaction between Hsp70 and Hsp90 in vivo, we identified Sti1 mutants that result in reduced recovery of Hsp70 in Hsp90 complexes. We also identified two Hsp90 mutants that exhibit a reduced Hsp70 interaction, which may help clarify the mechanism of client transfer between the two molecular chaperones.

scholarly journals Golgi-Localized, γ-Ear-Containing, Arf-Binding Protein Adaptors Mediate Insulin-Responsive Trafficking of Glucose Transporter 4 in 3T3-L1 Adipocytes

2005 ◽  
Vol 19 (8) ◽  
pp. 2145-2153 ◽  
Author(s):  
Lin V. Li ◽  
Konstantin V. Kandror
Keyword(s):  
Glucose Transporter ◽  
Adaptor Proteins ◽  
Dominant Negative ◽  
Glucose Transporter 4 ◽  
Glutathione S Transferase ◽  
Insulin Responsiveness ◽  
Vhs Domain

Abstract Small glucose transporter 4 (Glut4)-containing vesicles represent the major insulin-responsive compartment in fat and skeletal muscle cells. The molecular mechanism of their biogenesis is not yet elucidated. Here, we studied the role of the newly discovered family of monomeric adaptor proteins, GGA (Golgi-localized, γ-ear-containing, Arf-binding proteins), in the formation of small Glut4 vesicles and acquisition of insulin responsiveness in 3T3-L1 adipocytes. In these cells, all three GGA isoforms are expressed throughout the differentiation process. In particular, GGA2 is primarily present in trans-Golgi network and endosomes where it demonstrates a significant colocalization with the recycling pool of Glut4. Using the techniques of immunoadsorption as well as glutathione-S-transferase pull-down assay we found that Glut4 vesicles (but not Glut4 per se) interact with GGA via the Vps-27, Hrs, and STAM (VHS) domain. Moreover, a dominant negative GGA mutant inhibits formation of Glut4 vesicles in vitro. To study a possible role of GGA in Glut4 traffic in the living cell, we stably expressed a dominant negative GGA mutant in 3T3-L1 adipocytes. Formation of small insulin-responsive Glut4-containing vesicles and insulin-stimulated glucose uptake in these cells were markedly impaired. Thus, GGA adaptors participate in the formation of the insulin-responsive vesicular compartment from the intracellular donor membranes both in vivo and in vitro.

scholarly journals CRIg plays an essential role in intravascular clearance of bloodborne parasites by interacting with complement

2019 ◽  
Vol 116 (48) ◽  
pp. 24214-24220 ◽  
Author(s):  
Gongguan Liu ◽  
Yong Fu ◽  
Mohammed Yosri ◽  
Yanli Chen ◽  
Peng Sun ◽  
...  
Keyword(s):  
Essential Role ◽  
Parasitic Infection ◽  
Complement C3 ◽  
Critical Question ◽  
Independent Manner ◽  
African Trypanosomes ◽  
Recognition Receptor

Although CRIg was originally identified as a macrophage receptor for binding complement C3b/iC3b in vitro, recent studies reveal that CRIg functions as a pattern recognition receptor in vivo for Kupffer cells (KCs) to directly bind bacterial pathogens in a complement-independent manner. This raises the critical question of whether CRIg captures circulating pathogens through interactions with complement in vivo under flow conditions. Furthermore, the role of CRIg during parasitic infection is unknown. Taking advantage of intravital microscopy and using African trypanosomes as a model, we studied the role of CRIg in intravascular clearance of bloodborne parasites. Complement C3 is required for intravascular clearance of African trypanosomes by KCs, preventing the early mortality of infected mice. Moreover, antibodies are essential for complement-mediated capture of circulating parasites by KCs. Interestingly, reduced antibody production was observed in the absence of complement C3 during infection. We further demonstrate that CRIg but not CR3 is critically involved in KC-mediated capture of circulating parasites, accounting for parasitemia control and host survival. Of note, CRIg cannot directly catch circulating parasites and antibody-induced complement activation is indispensable for CRIg-mediated parasite capture. Thus, we provide evidence that CRIg, by interacting with complement in vivo, plays an essential role in intravascular clearance of bloodborne parasites. Targeting CRIg may be considered as a therapeutic strategy.

scholarly journals Zebrafish BID Exerts an Antibacterial Role by Negatively Regulating p53, but in a Caspase-8-Independent Manner

2021 ◽  
Vol 12 ◽  
Author(s):  
Zhitao Qi ◽  
Dong Yan ◽  
Lu Cao ◽  
Yang Xu ◽  
Mingxian Chang
Keyword(s):  
Bacterial Pathogen ◽  
P53 Gene ◽  
Bacterial Invasion ◽  
Caspase 8 ◽  
Pathogen Infection ◽  
Independent Manner ◽  
Invasion Assays

Bid (BH3-interacting domain death agonist), a member of the Bcl-2 family, plays a crucial role in the initiation of apoptosis. Independent of its apoptotic function, Bid is also involved in the regulation of inflammation and innate immunity. However, the role of Bid during bacterial pathogen infection remains unclear. In the present study, Bid of zebrafish (Dario rerio) was cloned and its functions during Edwardsiella ictaluri infection were investigated. Zebrafish Bid enhances the apoptosis rate of Epithelioma papulosum cyprini (EPC) cells following E. ictaluri infection. Importantly, in vitro and in vivo bacterial invasion assays showed that overexpressed Bid could significantly inhibit the invasion and proliferation of E. ictaluri. Real-time qPCR analysis revealed that p53 gene expression was downregulated in embryos microinjected with Bid-FLAG. Further, in vitro and in vivo bacterial invasion assays showed that overexpressed p53 increased the invasion and proliferation of E. ictaluri. Moreover, the invasion and proliferation of E. ictaluri were inhibited when co-overexpressing Bid and p53 in vivo and in vitro. Further, the numbers of E. ictaluri in larvae treated with Z-IETD-FMK (caspase-8 inhibitor) were higher than those of larvae without Z-IETD-FMK treatment, while the number of E. ictaluri in larvae microinjected with bid-Flag decreased significantly, even if the larvae were treated in advance with Z-IETD-FMK. Collectively, our study demonstrated a novel antibacterial activity of fish Bid, providing evidence for understanding the function of apoptosis associated gene in pathogen infection.

P13.17 CD95 gene silencing affects growth and invasiveness of glioma-initiating cells in a CD95L-independent manner

2021 ◽  
Vol 23 (Supplement_2) ◽  
pp. ii36-ii36
Author(s):  
C Quijano-Rubio ◽  
M Weller
Keyword(s):  
Gene Silencing ◽  
Apoptotic Cell ◽  
Gene Knockout ◽  
Acquired Resistance ◽  
Independent Manner ◽  
Glioma Initiating Cells ◽  
Induced Apoptosis

Abstract BACKGROUND CD95 (Fas/APO-1) holds a dual role of potential relevance in tumor development. CD95-CD95 ligand (CD95L) signaling regulates apoptotic cell death in CD95-expressing cells, but non-apoptotic, tumor-promoting CD95-CD95L signaling has been likewise described. Therapeutic stimulation of apoptotic CD95 signaling is associated with major clinical side effects. However, inhibition of tumor-promoting CD95 signaling may represent a promising treatment strategy for human cancers where potential tumor-promoting CD95 functions include invasiveness and cancer cell stemness, including glioblastoma. MATERIAL AND METHODS In this study, CD95 and CD95L expression was characterized in human glioma-initiating cells (GIC) in vitro and in vivo. CD95 and CD95L gene knockout (KO) GIC were generated by means of CRISPR-Cas9 and the effects of gene silencing were evaluated by assessing growth, clonogenicity, invasiveness and tumorigenicity in nude mice. RESULTS CD95 expression and sensitivity to exogenous CD95L-induced apoptosis were confirmed in selected GIC in vitro. CD95L expression was not detected. Upon CD95 KO, all GIC acquired resistance to CD95L-induced apoptosis. Furthermore, despite the confirmed absence of CD95L expression in vitro, CD95 KO S-24 GIC revealed decreased cell growth, inferior sphere forming capacity and decreased invasiveness. These data suggested a CD95L-independent tumor-promoting role of CD95 in S-24 GIC. In vivo, however, CD95 KO did not prolong the survival of glioma-bearing mice. Analyses of further GIC models are ongoing. CONCLUSION These data demonstrate that, unlike CD95, CD95L is not expressed in cultured human GIC and that CD95-CD95L interactions are not required for tumor-promoting CD95 signaling. Although CD95 KO is detrimental for S-24 GIC in vitro, CD95 KO alone does not affect survival in S-24 human GIC xenograft-bearing mice.

scholarly journals C-type natriuretic peptide enhances mouse preantral follicle growth

Reproduction ◽  
2019 ◽  
Vol 157 (5) ◽  
pp. 445-455 ◽  
Author(s):  
Guangyin Xi ◽  
Wenjing Wang ◽  
Sarfaraz A Fazlani ◽  
Fusheng Yao ◽  
Mingyao Yang ◽  
...  
Keyword(s):  
Natriuretic Peptide ◽  
Mechanistic Study ◽  
Preantral Follicle ◽  
Follicle Growth ◽  
Independent Manner ◽  
Preantral Follicles ◽  
Estrogen Synthesis

Compared to ovarian antral follicle development, the mechanism underlying preantral follicle growth has not been well documented. Although C-type natriuretic peptide (CNP) involvement in preantral folliculogenesis has been explored, its detailed role has not been fully defined. Here, we used mouse preantral follicles and granulosa cells (GCs) as a model for investigating the dynamic expression of CNP and natriuretic peptide receptor 2 (NPR2) during preantral folliculogenesis, the regulatory role of oocyte-derived growth factors (ODGFs) in natriuretic peptide type C (Nppc) and Npr2 expression, and the effect of CNP on preantral GC viability. Both mRNA and protein levels of Nppc and Npr2 were gradually activated during preantral folliculogenesis. CNP supplementation in culture medium significantly promoted the growth of in vitro-cultured preantral follicles and enhanced the viability of cultured GCs in a follicle-stimulating hormone (FSH)-independent manner. Using adult and prepubertal mice as an in vivo model, CNP pre-treatment via intraperitoneal injection before conventional superovulation also had a beneficial effect on promoting the ovulation rate. Furthermore, ODGFs enhanced Nppc and Npr2 expression in the in vitro-cultured preantral follicles and GCs. Mechanistic study demonstrated that the regulation of WNT signaling and estrogen synthesis may be implicated in the promoting role of CNP in preantral folliculogenesis. This study not only proves that CNP is a critical regulator of preantral follicle growth, but also provides new insight in understanding the crosstalk between oocytes and somatic cells during early folliculogenesis.

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