scholarly journals Involvement of Bacillus subtilis ClpE in CtsR Degradation and Protein Quality Control

2006 ◽  
Vol 188 (13) ◽  
pp. 4610-4619 ◽  
Author(s):  
Marcus Miethke ◽  
Michael Hecker ◽  
Ulf Gerth
Keyword(s):  
Bacillus Subtilis ◽  
Heat Stress ◽  
Protein Quality ◽  
Physiological Role ◽  
Structural Element ◽  
Wide Range ◽  
Adaptation Processes

ABSTRACT The heat-inducible CtsR regulon of Bacillus subtilis codes for three Clp proteins with chaperone or protease activity. While the importance of ClpC and ClpP has been elucidated for a wide range of cellular adaptation processes, this study deals with the physiological role of B. subtilis ClpE. Northern experiments and reporter gene analyses revealed that ClpE is essential both for efficient CtsR-dependent gene derepression and for rerepression during heat stress. ClpEP was found to destabilize the global regulator CtsR after heat shock in vivo with different kinetics than ClpCP, which is known to degrade CtsR in vitro and in vivo upon heat stress. Furthermore, ClpE was localized at heat-generated inclusion bodies by electron microscopy. The comparison of radiolabeled aggregated protein fractions of wild-type and clpE mutant cells during heat stress displayed a significant delay of protein disaggregation in the absence of ClpE. A kinetic Western blotting approach confirmed the long-term residence of ClpE in the insoluble cell fraction rather than in the cytoplasmic fraction. These observations indicate the involvement of ClpE in global protein disaggregation. As a characteristic structural element of ClpE, the N-terminal zinc finger domain was proven to be essential for basal in vitro ATPase activity.

Correlation between magnesium and potassium contents in muscle: role of Na(+)-K+ pump

1993 ◽  
Vol 264 (2) ◽  
pp. C457-C463 ◽  
Author(s):  
I. Dorup ◽  
T. Clausen
Keyword(s):  
Extensor Digitorum Longus ◽  
Extensor Digitorum ◽  
Deficient Diet ◽  
Young Rats ◽  
Wide Range ◽  
86Rb Influx

In young rats fed a Mg(2+)-deficient diet for 3 wk, Mg2+ and K+ contents in soleus and extensor digitorum longus muscles were significantly reduced and closely correlated. In isolated soleus muscles, Mg2+ depletion induced an even more pronounced loss of K+, and Mg2+ and K+ contents were correlated over a wide range (r = 0.95, P < 0.001). Extracellular Mg2+ (0-1.2 mM) caused no change in total or ouabain-suppressible 86Rb influx. After long-term incubation in Ca(2+)-Mg(2+)-free buffer with EDTA and EGTA, cellular Mg2+ and K+ contents were reduced by 35 and 15%, respectively, without any reduction in ATP and total or ouabain-suppressible 86Rb influx. In Mg(2+)-depleted muscles 42K efflux was increased by up to 42%, and repletion with Mg2+ produced a graded decrease. We conclude that Mg2+ and K+ contents are closely correlated in muscles Mg2+ depleted in vivo or in vitro and that neither extracellular nor moderate intracellular Mg2+ depletion affects total or Na(+)-K+ pump-mediated K+ influx. The reduced K+ content may rather be related to increased K+ efflux from the muscles.

scholarly journals Biogenesis, Biologic Function and Clinical Potential of Exosomes in Different Diseases

2020 ◽  
Vol 10 (13) ◽  
pp. 4428 ◽  
Author(s):  
Amany Magdy Beshbishy ◽  
Saad Alghamdi ◽  
ThankGod E. Onyiche ◽  
Muhammad Zahoor ◽  
Nallely Rivero-Perez ◽  
...  
Keyword(s):  
Soft Tissues ◽  
Protein Quality ◽  
Therapeutic Effects ◽  
Exosome Biogenesis ◽  
Wide Range ◽  
Potential Therapeutic Application ◽  
Intercellular Movement ◽  
Clinical Potential

Exosomes are extracellular vesicles (EVs) belonging to the nanovesicles family that function as signaling molecules between cells. After their first description in the late 1960s, interest in their potential as a research target has steadily increased. They are small secreted organelles with a single membrane that are well enriched in lipids, proteins, nucleic acids, and glycoconjugates. Exosomes take part in a larger communication network in which cells communicate between one another by DNA shuttling, proteins, RNA, and membrane-bound factors. The machinery of protein quality control occurs through the process termed “exosome biogenesis”. Furthermore, the pathway involved in intercellular movement of vesicles is vital in various aspects of human health and diseases. Due to their inherent properties, exosomes are currently being developed as potential therapeutic agents in a wide range of diseases including infectious and non-infectious diseases. Exosomes and other EVs sourced from Mesenchymal stem cells (MSCs) have been shown in different studies to possess therapeutic effects in diverse disease models either in vivo or in vitro. Some mechanisms and/or pathways that MSC-derived exosomes use to illustrate their therapeutic effect against some diseases have also been summarized. This review aims to highlight the recent findings and potential therapeutic application of exosomes in different diseases such as autoimmune, cardiovascular, obesity, neural, soft tissues, bone, and cartilage.

scholarly journals Role of Spore Coat Proteins in the Resistance of Bacillus subtilis Spores to Caenorhabditis elegans Predation

2008 ◽  
Vol 190 (18) ◽  
pp. 6197-6203 ◽  
Author(s):  
Maria-Halima Laaberki ◽  
Jonathan Dworkin
Keyword(s):  
Bacillus Subtilis ◽  
Caenorhabditis Elegans ◽  
Spore Coat ◽  
Coat Proteins ◽  
Host Interaction ◽  
Wide Range ◽  
In The Wild

ABSTRACT Bacterial spores are resistant to a wide range of chemical and physical insults that are normally lethal for the vegetative form of the bacterium. While the integrity of the protein coat of the spore is crucial for spore survival in vitro, far less is known about how the coat provides protection in vivo against predation by ecologically relevant hosts. In particular, assays had characterized the in vitro resistance of spores to peptidoglycan-hydrolyzing enzymes like lysozyme that are also important effectors of innate immunity in a wide variety of hosts. Here, we use the bacteriovorous nematode Caenorhabditis elegans, a likely predator of Bacillus spores in the wild, to characterize the role of the spore coat in an ecologically relevant spore-host interaction. We found that ingested wild-type Bacillus subtilis spores were resistant to worm digestion, whereas vegetative forms of the bacterium were efficiently digested by the nematode. Using B. subtilis strains carrying mutations in spore coat genes, we observed a correlation between the degree of alteration of the spore coat assembly and the susceptibility to the worm degradation. Surprisingly, we found that the spores that were resistant to lysozyme in vitro can be sensitive to C. elegans digestion depending on the extent of the spore coat structure modifications.

scholarly journals ILF and vReD pathways cooperate to control lysosomal transporter protein lifetimes

2017 ◽  
Author(s):  
Erin Kate McNally ◽  
Christopher Brett
Keyword(s):  
Quality Control ◽  
Heat Stress ◽  
Protein Quality ◽  
Control Individual ◽  
Degradation Pathways ◽  
Transporter Protein ◽  
Nutrient Transporter ◽  
Client Proteins

Lysosomal nutrient transporter proteins move lumenal products of biomaterial catabolism to the cytoplasm for reuse by the cell. Two mechanisms control their lifetimes: the ILF (IntraLumenal Fragment) and vReD (Vacuole REcycling and Degradation) pathways. But it is not clear if they function independently. Using S. cerevisiae as a model, here we show that the ILF pathway mediates constitutive turnover of the lysine transporter Ypq1 and zinc transporter Cot1-known vReD client proteins-in vivo and in vitro. In contrast, the vReD pathway mediates constitutive degradation of the amino acid transporter Vba4. Activation of TOR with cycloheximide enhances their degradation by these pathways. However, misfolding by heat stress shunts all three into the ILF pathway. Thus, both pathways control individual transporter lifetimes, although only the ILF pathway mediates protein quality control. The pathway chosen depends on protein fate: degradation is imminent by the ILF pathway, whereas the vReD pathway permits reuse.

scholarly journals Evidence that mammalian phosphatidylinositol transfer protein regulates phosphatidylcholine metabolism

1998 ◽  
Vol 335 (1) ◽  
pp. 175-179 ◽  
Author(s):  
Marie E. MONACO ◽  
Richard J. ALEXANDER ◽  
Gerry T. SNOEK ◽  
Nancy H. MOLDOVER ◽  
Karel W. A. WIRTZ ◽  
...  
Keyword(s):  
Physiological Role ◽  
Fold Increase ◽  
Transfer Protein ◽  
Antisense Mrna ◽  
Cell Clones ◽  
Phosphatidylinositol Transfer

Phosphatidylinositol transfer proteins (PITPs) and their yeast counterpart (SEC14p) possess the ability to bind phosphatidylinositol (PtdIns) and transfer it between membranes in vitro. However, the biochemical function of these proteins in vivo is unclear. In the present study, the physiological role of PITP was investigated by determining the biochemical consequences of lowering the cellular content of this protein. WRK-1 rat mammary tumour cells were transfected with a plasmid containing a full-length rat PITPα cDNA inserted in the antisense orientation and the resultant cell clones were analysed. Three clones expressing antisense mRNA for PITPα were compared with three clones transfected with the expression vector lacking the insert. The three antisense clones had an average of 25% less PITPα protein than control clones. Two of the three antisense clones also exhibited a decreased rate of growth. All three antisense clones exhibited a significant decrease in the incorporation of labelled precursors into PtdCho during a 90-min incubation period. Under the same conditions, however, there was no change in precursor incorporation into PtdIns. Further experimentation indicated that the decrease in precursor incorporation seen in antisense clones was not due to an increased rate of turnover. When choline metabolism was analysed more extensively in one control (2-5) and one antisense (4-B) clone using equilibrium-labelling conditions (48 h of incubation), the following were observed: (1) the decrease in radioactive labelling of PtdCho seen in short-term experiments was also observed in long-term experiments, suggesting that the total amount of PtdCho was lower in antisense-transfected clones (this was confirmed by mass measurements); (2) a similar decrease was seen in cellular sphingomyelin, lysoPtdCho and glycerophosphorylcholine; (3) an average two-fold increase in cellular phosphorylcholine was observed in the antisense-transfected clone; (4) cellular choline was, on average, decreased; and (5) cellular CDPcholine was not significantly altered.

scholarly journals Natural Variability in S-Adenosylmethionine (SAM)-Dependent Riboswitches: S-Box Elements in Bacillus subtilis Exhibit Differential Sensitivity to SAM In Vivo and In Vitro

2007 ◽  
Vol 190 (3) ◽  
pp. 823-833 ◽  
Author(s):  
Jerneja Tomšič ◽  
Brooke A. McDaniel ◽  
Frank J. Grundy ◽  
Tina M. Henkin
Keyword(s):  
Bacillus Subtilis ◽  
Structural Rearrangement ◽  
Natural Variability ◽  
Differential Sensitivity ◽  
In Vitro Assays ◽  
Leader Region ◽  
Structural Elements ◽  
Wide Range

ABSTRACT Riboswitches are regulatory systems in which changes in structural elements in the 5′ region of the nascent RNA transcript (the “leader region”) control expression of the downstream coding sequence in response to a regulatory signal in the absence of a trans-acting protein factor. The S-box riboswitch, found primarily in low-G+C gram-positive bacteria, is the paradigm for riboswitches that sense S-adenosylmethionine (SAM). Genes in the S-box family are involved in methionine metabolism, and their expression is induced in response to starvation for methionine. S-box genes exhibit conserved primary sequence and secondary structural elements in their leader regions. We previously demonstrated that SAM binds directly to S-box leader RNA, causing a structural rearrangement that results in premature termination of transcription at S-box leader region terminators. S-box genes have a variety of physiological roles, and natural variability in S-box structure and regulatory response could provide additional insight into the role of conserved S-box leader elements in SAM-directed transcription termination. In the current study, in vivo and in vitro assays were employed to analyze the differential regulation of S-box genes in response to SAM. A wide range of responses to SAM were observed for the 11 S-box-regulated transcriptional units in Bacillus subtilis, demonstrating that S-box riboswitches can be calibrated to different physiological requirements.

scholarly journals Intestinal absorption of linoleic acid in experimental renal failure

1991 ◽  
Vol 66 (3) ◽  
pp. 467-477
Author(s):  
M. V. Pahl ◽  
A. Barbari ◽  
N. D. Vaziri ◽  
D. Hollander ◽  
M. Yazdani ◽  
...  
Keyword(s):  
Renal Failure ◽  
Linoleic Acid ◽  
Water Layer ◽  
Short Term ◽  
Unstirred Water Layer ◽  
Wide Range ◽  
Significant Difference

Linoleic acid (LA) transport in rats with experimental short-term and long-term renal failure (RF) was compared with that of sham-operated normal animals on liberal food intake and pair-fed animals. The perfusions in vivo and incubations in vitro were conducted using a micellar solution containing a wide range of LA concentrations. Both absorption in vivo and uptake in vitro of LA were significantly reduced in animals with short-term RF. Lipid extraction and separation by thin-layer chromatography revealed a marked LA trapping as trilinolein (TL) in the perfused intestinal tissue in the short-term RF group. The esterification process, as defined by the rate of LA incorporation into TL, was moderately reduced in short-term RF animals. The thickness of the unstirred water layer showed no significant difference among the groups studied. In contrast, animals with long-term RF exhibited normal absorption of LA in vivo at all concentrations tested. In conclusion, LA absorption is reduced in short-term RF and restored in long-term RF. Several steps including LA transport into and TL transport out of the enterocyte and the esterification process were impaired in short-term RF. These changes are not due to alteration in the unstirred water layer, anorexia, weight loss or a rapid effect of uraemic chemical environment or circulatory factors.

A muscle-powered energy delivery system and means for chronic in vivo testing

1999 ◽  
Vol 86 (6) ◽  
pp. 2106-2114 ◽  
Author(s):  
Dennis R. Trumble ◽  
James A. Magovern
Keyword(s):  
In Vitro Tests ◽  
Access Port ◽  
Wide Range ◽  
Piston Displacement ◽  
In Vivo Testing ◽  
Implant Testing

Electrically stimulated skeletal muscle represents a potentially unlimited source of energy for the actuation of motor prostheses. Devices to harvest and deliver contractile power have proven mechanically feasible, but long-term efficacy has not been demonstrated. This report describes recent refinements in muscle energy converter (MEC) design and details the development of an implantable afterload chamber (IAC) designed to facilitate implant testing. The IAC comprises a fluid-filled bladder housed within a titanium cylinder that connects directly to the MEC. A vascular access port allows percutaneous measurement and adjustment of air pressure within the housing and provides a means both to monitor MEC function and to control hydraulic loading conditions. Data from in vitro tests show that IAC pressure mirrors changes in MEC-piston displacement over a wide range of actuation speeds and stroke lengths. Stroke lengths and actuation forces calculated from IAC pressure readings were typically found to be within 5% of measured values. This testing scheme may yield important information in regard to the ability to harness energy from in situ muscle over prolonged periods.

scholarly journals Cyclic-di-GMP is required for corneal infection byPseudomonas aeruginosaand modulates host immunity

2017 ◽  
Author(s):  
Joey Kuok Hoong Yam ◽  
Thet Tun Aung ◽  
Song Lin Chua ◽  
Yingying Cheng ◽  
Gurjeet Singh Kohli ◽  
...  
Keyword(s):  
Extracellular Polymeric Substances ◽  
Antimicrobial Treatment ◽  
Guanosine Monophosphate ◽  
Rna Seq ◽  
Physiological Characterization ◽  
Wide Range

AbstractBiofilms are extremely tolerant toward antimicrobial treatment and host immune clearance due to their distinct physiology and protection by extracellular polymeric substances. Bis-(3´-5´)-cyclic dimeric guanosine monophosphate (c-di-GMP) is an essential messenger that regulates biofilm formation by a wide range of bacteria. However, there is a lack of physiological characterization of biofilmsin vivoas well as the roles of c-di-GMP signaling in mediating host-biofilm interactions. Here, we employed dual RNA-Seq to characterize the host and pathogen transcriptomes duringPseudomonas aeruginosainfection using a mouse keratitis model.In vivo P. aeruginosabiofilms maintained a distinct physiology compared within vitro P. aeruginosabiofilms, with enhanced virulence and iron uptake capacity. C-di-GMP synthesis was enhanced inP. aeruginosacellsin vivo,potentially due to down-regulation of the expression of several phosphodiesterases (e.g., DipA, NbdA). Increased intracellular c-di-GMP levels were required for long-term ocular colonization ofP. aeruginosaand impaired host innate immunity.

Evaluation of 99mTc-Label led Vitamin K4 as an Agent for Testicular Imaging

1991 ◽  
Vol 30 (01) ◽  
pp. 35-39 ◽  
Author(s):  
H. S. Durak ◽  
M. Kitapgi ◽  
B. E. Caner ◽  
R. Senekowitsch ◽  
M. T. Ercan
Keyword(s):  
Soft Tissue ◽  
Room Temperature ◽  
Normal Subjects ◽  
Left Testis ◽  
Wide Range ◽  
Activity Ratios ◽  
The Right ◽  
Radioactivity Levels

Vitamin K4 was labelled with 99mTc with an efficiency higher than 97%. The compound was stable up to 24 h at room temperature, and its biodistribution in NMRI mice indicated its in vivo stability. Blood radioactivity levels were high over a wide range. 10% of the injected activity remained in blood after 24 h. Excretion was mostly via kidneys. Only the liver and kidneys concentrated appreciable amounts of radioactivity. Testis/soft tissue ratios were 1.4 and 1.57 at 6 and 24 h, respectively. Testis/blood ratios were lower than 1. In vitro studies with mouse blood indicated that 33.9 ±9.6% of the radioactivity was associated with RBCs; it was washed out almost completely with saline. Protein binding was 28.7 ±6.3% as determined by TCA precipitation. Blood clearance of 99mTc-l<4 in normal subjects showed a slow decrease of radioactivity, reaching a plateau after 16 h at 20% of the injected activity. In scintigraphic images in men the testes could be well visualized. The right/left testis ratio was 1.08 ±0.13. Testis/soft tissue and testis/blood activity ratios were highest at 3 h. These ratios were higher than those obtained with pertechnetate at 20 min post injection.99mTc-l<4 appears to be a promising radiopharmaceutical for the scintigraphic visualization of testes.

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