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Robert H. Smith Faculty of Agriculture, Food and Environment,
The Hebrew University of Jerusalem, 
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Vetvicka, V. ; Gover, G. ; Hayby, H. ; Danay, O. ; Ezov, N. ; Hadar, Y. ; Schwartz, B. Immunomodulating effects exerted by glucans extracted from the king oyster culinary-medicinal mushroom pleurotus eryngii (agaricomycetes) grown in substrates containing various concentrations of olive mill waste. International Journal of Medicinal Mushrooms 2019, 21, 765-781. Publisher's VersionAbstract
We have recently demonstrated that we could enhance glucan content in Pleurotus eryngii following cultivation of the mushrooms on a substrate containing different concentrations of olive mill solid waste (OMSW). These changes are directly related to the content of OMSW in the growing substrate. Using dextran sulfate sodium (DSS)-inflammatory bowel disease (IBD) mice model, we measured the colonic inflammatory response to the different glucan preparations. We found that the histology damaging score (HDS) resulting from DSS treatment reach a value of 11.8 ± 2.3 were efficiently downregulated by treatment with the fungal extracted glucans. Glucans extracted from stalks cultivated at 20% OMSW downregulated to a HDS value of 6.4 ± 0.5 whereas those cultivated at 80% OMSW showed the strongest effects (5.5 ± 0.6). Similar downregulatory effects were obtained for expression of various intestinal cytokines. All tested glucans were equally effective in regulating the number of CD14/CD16 monocytes from 18.2 ± 2.7% for DSS to 6.4 ± 2.0 for DSS + glucans extracted from stalks cultivated at 50% OMSW. We tested the effect of glucans on lipopolysaccharide- induced production of TNF-α, which demonstrated that stalk-derived glucans were more effective than caps-derived glucans. Isolated glucans competed with anti-Dectin-1 and anti-CR3 antibodies, indicating that they contain β-glucans recognized by these receptors. In conclusion, the most effective glucans in ameliorating IBD-associated symptoms induced by DSS treatment in mice were glucan extracts prepared from the stalk of P. eryngii grown at higher concentrations of OMSW. We conclude that these stress-induced growing conditions may be helpful in selecting more effective glucans derived from edible mushrooms. © 2019 by Begell House, Inc.
Israeli, E. ; Adler Berken, N. ; Gover, O. ; Waechtershaeuser, E. ; Graeve, L. ; Schwartz, B. Recombinant ostreolysin (rOly) inhibits the anti-adipogenic Hedgehog (Hh) signaling pathway in 3T3-L1 cells. Journal of Functional Foods 2019, 59, 185-193. Publisher's VersionAbstract
Obesity is a nutrition-associated disorder result of an imbalance between energy intake and energy expenditure. Changing adipocytes differentiation patterns is considered as a strategy to treat obesity-related disorders. Recently, much interest is focused on the role of posttranslational modifications of tubulin on adipocyte differentiation. We recently demonstrated that a recombinant version of the fungal protein Ostreolysin (rOly) drastically affects metabolism of adipose tissue. The aim of the present study is to extend our understanding of the in vitro effects of rOly on different adipocytes. We demonstrate that rOly inhibits the anti-adipogenic Hedgehog (Hh) signaling pathway in 3T3-L1 cells. Additionally, rOly affected the gene expression levels of SQSTM1 and Collagen type 1, which are mediated by AMP-activated protein kinase (AMPK) activity in 3T3-L1 cells. We provide a potential molecular mechanistic approach describing that the effect of rOly on adipocytes is mediated by tubulin acetylation and AMPK phosphorylation. © 2019 Elsevier Ltd
Miron, N. ; Tirosh, O. Cholesterol Prevents Hypoxia-Induced Hypoglycemia by Regulation of a Metabolic Ketogenic Shift. Oxidative Medicine and Cellular Longevity 2019, 2019. Publisher's VersionAbstract
Blood cholesterol levels have been connected to high-altitude adaptation. In the present study, we treated mice with high-cholesterol diets following exposure to acute hypoxic stress and evaluated the effects of the diets on whole-body, liver glucose, and liver fat metabolism. For rapid cholesterol liver uptake, 6-week-old male C57BL/J6 mice were fed with high-cholesterol/cholic acid (CH) diet for 6 weeks and then were exposed to gradual oxygen level reduction for 1 h and hypoxia at 7% oxygen for additional 1 hour using a hypoxic chamber. Animals were than sacrificed, and metabolic markers were evaluated. Hypoxic treatment had a strong hypoglycemic effect that was completely blunted by CH treatment. Decreases in gluconeogenesis and glycogenolysis as well as an increase in ketone body formation were observed. Such changes indicate a metabolic shift from glucose to fat utilization due to activation of the inducible nitric oxide synthase/AMPK axis in the CH-treated animals. Increased ketogenesis was also observed in vitro in hepatocytes after cholesterol treatment. In conclusion, our results show for the first time that cholesterol contributes to metabolic shift and adaptation to hypoxia in vivo and in vitro through induction of HIF-1α and iNOS expression. © 2019 Naama Miron and Oren Tirosh.
Anavi, S. ; Tirosh, O. iNOS as a metabolic enzyme under stress conditions. Free Radical Biology and Medicine 2019. Publisher's VersionAbstract
Nitric oxide (NO) is a free radical acting as a cellular signaling molecule in many different biochemical processes. NO is synthesized from L-arginine through the action of the nitric oxide synthase (NOS) family of enzymes, which includes three isoforms: endothelial NOS (eNOS), neuronal NOS (nNOS) and inducible NOS (iNOS). iNOS-derived NO has been associated with the pathogenesis and progression of several diseases, including liver diseases, insulin resistance, obesity and diseases of the cardiovascular system. However, transient NO production can modulate metabolism to survive and cope with stress conditions. Accumulating evidence strongly imply that iNOS-derived NO plays a central role in the regulation of several biochemical pathways and energy metabolism including glucose and lipid metabolism during inflammatory conditions. This review summarizes current evidence for the regulation of glucose and lipid metabolism by iNOS during inflammation, and argues for the role of iNOS as a metabolic enzyme in immune and non-immune cells. © 2019
Kanner, J. ; Shpaizer, A. ; Nelgas, L. ; Tirosh, O. S-Nitroso- N-acetylcysteine (NAC-SNO) as an Antioxidant in Cured Meat and Stomach Medium. Journal of Agricultural and Food Chemistry 2019, 67, 10930-10936. Publisher's VersionAbstract
The stability of lipids in meat products depends on the initial concentration of hydroperoxides, the catalytic involvement of metal ions and myoglobin, endogenous antioxidants, and biological and technological factors. Ground meat was treated with additives, sealed in vacuum bags, heated to 75 °C, and stored opened to air at 4 °C. S-Nitroso-N-acetylcysteine (NAC-SNO) at concentration like nitrite used by the industry prevents lipid peroxidation in the product, even after storage for 1 month at 4 °C. The same simulated treatments at different concentrations of both compounds show that NAC-SNO acts as an antioxidant ∼4-fold better than nitrite at pH 6.2 or 3.0. Ascorbic acid significantly improves nitrite antioxidant effect. NAC-SNO was found to prevent, much better than nitrite, accumulation of reactive aldehydes and hydroxynonenal protein modification. In condition like those used by the industry for meat products processing, NAC-SNO acts better than nitrite to provide antioxidant protection without the side effect of N-nitrosation, oxidation, and the loss of nutrient generated by nitrite. © 2019 American Chemical Society.
Alber, J. ; Alladi, S. ; Bae, H. - J. ; Barton, D. A. ; Beckett, L. A. ; Bell, J. M. ; Berman, S. E. ; Biessels, G. J. ; Black, S. E. ; Bos, I. ; et al. White matter hyperintensities in vascular contributions to cognitive impairment and dementia (VCID): Knowledge gaps and opportunities. Alzheimer's and Dementia: Translational Research and Clinical Interventions 2019, 5 107-117. Publisher's VersionAbstract
White matter hyperintensities (WMHs) are frequently seen on brain magnetic resonance imaging scans of older people. Usually interpreted clinically as a surrogate for cerebral small vessel disease, WMHs are associated with increased likelihood of cognitive impairment and dementia (including Alzheimer's disease [AD]). WMHs are also seen in cognitively healthy people. In this collaboration of academic, clinical, and pharmaceutical industry perspectives, we identify outstanding questions about WMHs and their relation to cognition, dementia, and AD. What molecular and cellular changes underlie WMHs? What are the neuropathological correlates of WMHs? To what extent are demyelination and inflammation present? Is it helpful to subdivide into periventricular and subcortical WMHs? What do WMHs signify in people diagnosed with AD? What are the risk factors for developing WMHs? What preventive and therapeutic strategies target WMHs? Answering these questions will improve prevention and treatment of WMHs and dementia. © 2019 The Authors
Swan, W. I. ; Pertel, D. G. ; Hotson, B. ; Lloyd, L. ; Orrevall, Y. ; Trostler, N. ; Vivanti, A. ; Howarter, K. B. ; Papoutsakis, C. Nutrition Care Process (NCP) Update Part 2: Developing and Using the NCP Terminology to Demonstrate Efficacy of Nutrition Care and Related Outcomes. Journal of the Academy of Nutrition and Dietetics 2019, 119, 840-855. Publisher's Version
Perito, M. A. ; Sacchetti, G. ; Di Mattia, C. D. ; Chiodo, E. ; Pittia, P. ; Saguy, I. ; Cohen, E. Buy Local! Familiarity and Preferences for Extra Virgin Olive Oil of Italian Consumers. Journal of Food Products Marketing 2019, 25, 462-477. Publisher's VersionAbstract
Over the last few years, the origin of the local product has played a central role in consumer choices. This study explores what Italian consumers want and look for when purchasing olive oil by combining a web-based survey and a perceived analysis technique. In particular, preferences for different olive oil attributes as well as the psychographic traits of respondents were revealed through a web-based questionnaire administered to Italian consumers (N = 179). From this questionnaire, respondents who indicated their availability to participate further underwent a preference test under blind conditions (N = 99). Respondents also did an expectation test based on the visual observation of the labels. Results showed that the majority of consumers considered local production, PDO and region as factors of highest importance in determining olive oil quality. © 2019, © 2019 Taylor & Francis.
Achmon, Y. ; Dowdy, F. R. ; Simmons, C. W. ; Zohar-Perez, C. ; Rabinovitz, Z. ; Nussinovitch, A. Degradation and bioavailability of dried alginate hydrocolloid capsules in simulated soil system. Journal of Applied Polymer Science 2019, 136. Publisher's VersionAbstract
Hydrocolloid capsules are common chemical carriers used in many types of applications in foods, biotechnology, and agriculture. Alginate microbeads and macrobeads are some of the more prevalent types of hydrocolloid capsules. Most studies to date have focused on alginate carrier applications but only a few have looked at their bioavailability after use. In this study, alginate carriers were subjected to simulated field conditions and their biodegradation in the soil was evaluated by respiration measurements, visualization, and volatile solids reduction. Using respiration rate, the degradation rate was calculated at 32 ± 3.1% (w/w) after 2 months. The visually estimated volume and volatile solids reduction gave degradation rates of 40 ± 8.6% (v/v) and 22.5 ± 2.5% (w/w), respectively. Moreover, water-loss calculations suggested that the carriers can serve as a stand-alone soil amendment for water retention. These findings emphasize the importance of studying hydrocolloid bioavailability in the soil and alginate carrier suitability for future applications. © 2019 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2019, 136, 48142. © 2019 Wiley Periodicals, Inc.
Blaychfeld-Magnazi, M. ; Reshef, N. ; Zornitzki, T. ; Madar, Z. ; Knobler, H. The effect of a low-carbohydrate high-fat diet and ethnicity on daily glucose profile in type 2 diabetes determined by continuous glucose monitoring. European Journal of Nutrition 2019. Publisher's VersionAbstract
{Background and aims: Nutrition is an integral part of type 2 diabetes (T2DM) treatment, but the optimal macronutrient composition is still debated and previous studies have not addressed the role of ethnicity in dietary response. The current study aims were to compare the effect of short-term glycemic response to low-carbohydrate high-fat (LC-HF) diet vs. high-carbohydrate low-fat (HC-LF) diet using continuous glucose monitoring (CGM) and to evaluate the response of individuals with T2DM of Yemenite (Y-DM) and non-Yemenite origin (NY-DM). Methods: Twenty T2DM males, ten Y-DM and ten NY-DM underwent meal tolerance test and indexes of insulin resistance and secretion were calculated. Subsequently, patients were connected to CGM to assess daily glycemic control and glucose variability in response to isocaloric HC-LF or LC-HF diet, receiving each diet for 2 days by providing prepared meals. Daily glucose levels, area under the glucose curve (G-AUC) and parameters of glucose variability [standard deviation (SD), mean amplitude of glycemic excursions (MAGE) and mean absolute glucose (MAG)] were evaluated. Results: The LC-HF resulted in a significantly lower G-AUC (p < 0.001) and in lower variability parameters (p < 0.001) vs. the HC-LF diet. However, Y-DM showed less reduction in glucose variability indices upon diet-switching vs. NY-DM; MAGE decreased, respectively, by 69% vs. 89%
Nakonechny, F. ; Avisar, D. ; Ludmer, Z. ; Brauner, N. ; Ullmann, A. Application of Partially Miscible Solvent System for an Efficient Extraction of Organic Pollutants from Contaminated Sludge. Water, Air, and Soil Pollution 2019, 230. Publisher's VersionAbstract
A systematic study of extraction of various organic pollutants from highly contaminated solid media (e.g., sludge) by applying a phase transition of a benign partially miscible solvent system composed of water/ethyl acetate/ethanol was conducted. This solvent system possesses an upper critical solution temperature of about 55 °C. The efficiency of the phase transition extraction (PTE) process is found to be higher and much faster compared to those obtained without phase transition (at ambient temperature). The influence of various operating conditions on the process efficiency was investigated. The performance of the phase transition extraction when applied on contaminated sludge is much better than the extraction with ethyl acetate only, although the latter is shown to be a very efficient solvent for extracting various organic contaminants (e.g., pharmaceuticals, persistent organic pollutants) from aqueous solutions. The efficiency of phase transition extraction from the aqueous solution was somewhat lower than that achieved with ethyl acetate, but it shows a clear advantage in the presence of detergents, as emulsion formation is prevented. © 2019, Springer Nature Switzerland AG.
Dolev, N. ; Katz, Z. ; Ludmer, Z. ; Ullmann, A. ; Brauner, N. ; Goikhman, R. New insights into chelator recycling by a chelating resin: From molecular mechanisms to applicability. Chemosphere 2019, 215, 800-806. Publisher's VersionAbstract
As part of the project of developing a “green” and highly feasible soil remediation process, recycling an eco-friendly chelating agent, glycine, using Chelex-100 chelating resin, was studied. Two model complexes, copper and nickel glycinates, were tested under various conditions, including equivalent viscosity but different temperature conditions. Two similar complexes demonstrated very different reactivity towards Chelex-100. An in-depth study led to the discovery of unusual metal-dependent mechanisms of the complex-to-resin metal transfer. Particularly, nickel transfer proceeds via a dissociative mechanism, whereas copper transfer does not require pre-dissociation of the complexes, and proceeds via the associative ligand-exchange mechanism. Both processes result in the recovery of the used chelator. The glycine solution was applied on the spiked soil, then recovered on Chelex-100 resin and successfully reused, thus demonstrating a proof of the concept. These findings contribute to the science, strategies, and methodology of both water purification and chelator recycling fields. © 2018 Elsevier Ltd
Gertler, A. ; Solomon, G. Pegylated Human Leptin D23L Mutant - Preparation and biological activity in vitro and in vivo in male ob /ob mice. Endocrinology 2019, 160, 891-898. Publisher's VersionAbstract
Recombinant monomeric human leptin (hLEP) and its D23L mutant were prepared in Escherichia coli and pegylated at their N-terminus using 20-kDa methoxy pegylated (PEG)-propionylaldehyde. As determined by both SDS-PAGE and size-exclusion chromatography, the pegylated proteins consisted of >90% monopegylated and <10% double-pegylated species. Circular dichroism spectra showed that their secondary structure, characteristic of all four α-helix bundle cytokines, was not affected by either the D23L mutation or pegylation. Because of the D23L mutation, affinity for hLEP receptor increased 25- and 40-fold for the pegylated and nonpegylated mutant, respectively. However, whereas the proliferation-promoting activity in vitro of nonmutated and mutated nonpegylated hLEP was identical, that of the respective pegylated mutant was approximately sixfold higher compared with the pegylated nonmutated hLEP. This difference was also seen in vivo. Both pegylated hLEPs at all doses significantly decreased body weight and food consumption, as compared with the vehicle-treated control. Once-daily administration of pegylated hLEP D23L at doses of 0.1, 0.3, and 1 mg/kg for 14 consecutive days in ob/ob mice resulted in significantly decreased body weight and food consumption as compared with respective pegylated hLEP-treated animals, with the biggest difference observed at 0.1 mg/kg. Repeated administration of either pegylated hLEP D23L or pegylated hLEP significantly decreased blood glucose levels compared with the control before glucose challenge and after oral glucose tolerance test, but with no difference between the two treatments. The pegylated hLEP D23L mutant seems to be a more potent reagent suitable for in vivo studies than the pegylated nonmutated hLEP. © Copyright 2019 Endocrine Society.
Ramos-Nieves, J. M. ; Giesy, S. L. ; Schwark, W. S. ; Gertler, A. ; Boisclair, Y. R. Technical note: Effects of pegylation and route of administration on leptin kinetics in newborn lambs. Journal of Animal Science 2019, 97, 3768-3775. Publisher's VersionAbstract
Chronic energy insufficiency resulting from inadequate feed intake or increased nutrient demand reduces plasma leptin in ruminants. Treatment of energy-deficient ruminants with exogenous leptin has identified some physiological consequences of reduced plasma leptin, but their full complement remains unknown. Additional leptin-dependent responses could be identified by using strategies that interfere with leptin signaling such as administration of leptin mutants that act as competitive antagonists. The effectiveness of these antagonists depends on their fold excess over endogenous leptin, and this condition can be achieved under in vivo conditions by extending the half-life (t1/2) of the antagonist by addition of a polyethylene glycol (PEG) molecule (pegylation). Use of this approach in ruminants, however, is limited by the lack of information on the t1/2 of native and pegylated leptin and on the most effective route of administration. To answer these questions, newborn lambs (n = 3) were injected with an intravenous (i.v.) bolus of 150 µg of human leptin followed by blood sampling over the next 12 h. Analysis of the semilog plasma leptin concentration over time yielded a t1/2 of 43 ± 4.9 min; an i.v. bolus of 276 µg of bovine leptin yielded a comparable t1/2 (P > 0.05). Next, newborn lambs (n = 4) received a single dose of 229 μg/ kg of metabolic body weight (BW0.75) of pegylated super human leptin antagonist (PEG-SHLA) via the i.v. or subcutaneous (s.c.) route. Plasma PEG-SHLA concentration reached a peak of 1,528 ± 78 ng/mL after 1 min and a nadir of 71 ± 9 ng/mL after 24 h with the i.v. route versus a peak of 423 ± 43 ng/mL after 300 min and a nadir of 146 ± 22 ng/mL after 24 h for the s.c. route; the t1/2 of PEG-SHLA was 394 ± 29 min for the i.v. route and 433 ± 58 min for the s.c. route. Finally, plasma concentration of PEG-SHLA was modeled when given either i.v. or s.c. at a dose of 229 μg/kg BW0.75 every 12 h. Once a steady state was reached, peak and lowest concentrations PEG-SHLA over the 12-h windows were 2,269 and 403 ng/mL for the i.v. route and 814 and 555 ng/mL for the s.c. route. Weighted PEG-SHLA concentrations over the 12-h period were 1,455 and 713 ng/mL for the i.v. and s.c. route, translating into 364- and 178-fold excess over endogenous plasma leptin. These data confirm the effectiveness of pegylation in extending the t1/2 of leptin antagonists in newborn lambs and in increasing their circulation in fold excess over endogenous leptin. © The Author(s) 2019. Published by Oxford University Press on behalf of the American Society of Animal Science. All rights reserved.
Kohay, H. ; Bilkis, I. ; Mishael, Y. Effect of polycation charge density on polymer conformation at the clay surface and consequently on pharmaceutical binding. Journal of Colloid and Interface Science 2019, 552, 517-527. Publisher's VersionAbstract
Polycation conformation upon adsorption to a negatively charged surface can be modulated by its charge density. At high charge density monomers directly interact with the surface in a ‘trains’ conformation and as charge density decreases a high degree of monomers dangle into solution in a ‘loops and tails’ conformation. In this study, the conformations of polycations upon adsorption to montmorillonite, as a function of polycation charge (20, 50 and 100% of the monomers, denoted as P-Q20, P-Q50 and P-Q100) were characterized and in accordance with their conformation, the adsorption of non-ionic and anionic molecules by the composite was tested. As expected, the adsorption of the nonionic pharmaceuticals increased to a composite with a ‘loops and tails’ conformation, due to both conformation and chemical properties. On the other hand, the anionic molecules, gemfibrozil and diclofenac, preferably adsorbed to composites with higher charge density (Q50 or Q100 composites). However, they showed different tendency toward the composites, i.e. higher adsorption of diclofenac by Q100 composite vs. higher adsorption of gemfibrozil by Q50 composite. To elucidate the differences in adsorption between these two pharmaceuticals, density functional theory calculations were employed. Both gemfibrozil and diclofenac were found to be better stabilized by methyl pyridinium sites (prevail in Q100 composite). The preferable adsorption of gemfibrozil by Q50 composite was explained by the presence of ‘loops and tails’ conformation enabling additional adsorption sites and diverse monomer-target molecule orientations. © 2019
Thawabteh, A. ; Lelario, F. ; Scrano, L. ; Bufo, S. A. ; Nowak, S. ; Behrens, M. ; Di Pizio, A. ; Niv, M. Y. ; Karaman, R. Bitterless guaifenesin prodrugs—design, synthesis, characterization, in vitro kinetics, and bitterness studies. Chemical Biology & Drug Design 2019, 93, 262-271. Publisher's VersionAbstract
Abstract A respected number of drugs suffer from bitter taste which results in patient incompliance. With the aim of solving the bitterness of guaifenesin, dimethyl maleate, maleate, glutarate, succinate, and dimethyl succinate prodrugs were designed and synthesized. Molecular orbital methods were utilized for the design of the ester prodrugs. The density functional theory (DFT) calculations revealed that the hydrolysis efficiency of the synthesized prodrugs is significantly sensitive to the pattern of substitution on C=C bond and distance between the nucleophile and the electrophile. The hydrolysis of the prodrugs was largely affected by the pH of the medium. The experimental t1/2 for the hydrolysis of guaifenesin dimaleate ester prodrugs in 1N HCl was the least and for guaifenesin dimethyl succinate was the highest. Functional heterologous expression of TAS2R14, a broadly tuned bitter taste receptor responding to guaifenesin, and experiments using these prodrugs revealed that, while some of the prodrugs still activated the receptor similarly or even stronger than the parent substance, succinate derivatization resulted in the complete loss of receptor responses. The predicted binding modes of guaifenesin and its prodrugs to the TAS2R14 homology model suggest that the decreased activity of the succinate derivatives may be caused by a clash with Phe247.
Di Pizio, A. ; Ben Shoshan-Galeczki, Y. ; Hayes, J. E. ; Niv, M. Y. Bitter and sweet tasting molecules: It's complicated. Neuroscience Letters 2019, 700, 56 - 63. Publisher's VersionAbstract
“Bitter” and “sweet” are frequently framed in opposition, both functionally and metaphorically, in regard to affective responses, emotion, and nutrition. This oppositional relationship is complicated by the fact that some molecules are simultaneously bitter and sweet. In some cases, a small chemical modification, or a chirality switch, flips the taste from sweet to bitter. Molecules humans describe as bitter are recognized by a 25-member subfamily of class A G-protein coupled receptors (GPCRs) known as TAS2Rs. Molecules humans describe as sweet are recognized by a TAS1R2/TAS1R3 heterodimer of class C GPCRs. Here we characterize the chemical space of bitter and sweet molecules: the majority of bitter compounds show higher hydrophobicity compared to sweet compounds, while sweet molecules have a wider range of sizes. Importantly, recent evidence indicates that TAS1Rs and TAS2Rs are not limited to the oral cavity; moreover, some bitterants are pharmacologically promiscuous, with the hERG potassium channel, cytochrome P450 enzymes, and carbonic anhydrases as common off-targets. Further focus on polypharmacology may unravel new physiological roles for tastant molecules.
Kolitsida, P. ; Zhou, J. ; Rackiewicz, M. ; Nolic, V. ; Dengjel, J. ; Abeliovich, H. Phosphorylation of mitochondrial matrix proteins regulates their selective mitophagic degradation. Proceedings of the National Academy of Sciences 2019. Publisher's VersionAbstract
Mitochondrial dysfunction underlies many age-related human pathologies. In normal cells, defective mitochondria are often degraded by mitophagy, a process in which these mitochondria are engulfed in autophagosomes and sent for degradation in the lysosome/vacuole. Surprisingly, studies on mitophagy in diverse eukaryotic organisms reveal an unexpected dimension of protein-level selectivity, wherein individual protein species exhibit divergent rates of mitophagic degradation. In this report, we show that this surprising intramitochondrial selectivity can be generated by differential phosphorylation of individual mitochondrial protein species, and we identify mitochondrial phosphatases and kinases that contribute to this regulation. By identifying a mechanism that regulates the intramitochondrial selectivity of mitophagic degradation, our findings open the door to potential manipulation of the quality-control process in the future.Mitophagy is an important quality-control mechanism in eukaryotic cells, and defects in mitophagy correlate with aging phenomena and neurodegenerative disorders. It is known that different mitochondrial matrix proteins undergo mitophagy with very different rates but, to date, the mechanism underlying this selectivity at the individual protein level has remained obscure. We now present evidence indicating that protein phosphorylation within the mitochondrial matrix plays a mechanistic role in regulating selective mitophagic degradation in yeast via involvement of the Aup1 mitochondrial protein phosphatase, as well as 2 known matrix-localized protein kinases, Pkp1 and Pkp2. By focusing on a specific matrix phosphoprotein reporter, we also demonstrate that phospho-mimetic and nonphosphorylatable point mutations at known phosphosites in the reporter increased or decreased its tendency to undergo mitophagy. Finally, we show that phosphorylation of the reporter protein is dynamically regulated during mitophagy in an Aup1-dependent manner. Our results indicate that structural determinants on a mitochondrial matrix protein can govern its mitophagic fate, and that protein phosphorylation regulates these determinants.
Wang, S. ; Duan, Y. ; Yan, Y. ; Adar, C. ; Braslavsky, I. ; Chen, B. ; Huang, T. ; Qiu, S. ; Li, X. ; Inglis, B. M. ; et al. Improvement of sperm cryo-survival of cynomolgus macaque (Macaca fascicularis) by commercial egg-yolk–free freezing medium with type III antifreeze protein. Animal Reproduction Science 2019, 210, 106177. Publisher's VersionAbstract
When nonhuman primate sperm undergoes cryopreservation in an egg yolk medium there is an increased risk that the egg yolk might adversely affect the sperm due to containing of avian pathogens. Although commercial egg-yolk-free medium for human sperm cryopreservation has been used for macaque sperm, the cryo-survival remains less than optimal. The present study, therefore, was conducted to determine the optimal concentration of antifreeze protein (AFP) III supplemented in a commercial egg-yolk-free medium for cynomolgus macaque (Macaca fascicularis) sperm cryo-survival. The function of frozen-thawed sperm was evaluated by post-thaw sperm motility, acrosome integrity, and mitochondrial function. Results indicate that the sperm motilities were greater when 0.1, 1, and 10 μg/ml of AFP III were supplemented into the sperm freezing medium (P < 0.05). In addition, the mitochondrial membrane potential was greater in the sperm cryopreserved with the medium that was supplemented with 0.1 μg/ml of AFP III (P < 0.05). The addition of AFP III at any of the concentrations, however, did not have any cryoprotection effect on the sperm acrosome, and the greatest concentrations of AFP III at 100 and 200 μg/ml had detrimental effects on acrosomal integrity (P < 0.05). Results of the present study indicated the methods used are effective for the cryopreservation of cynomolgus monkey sperm while reducing associated health risks due to avian pathogens being present in egg yolk-based extenders.
Yehuda, A. ; Slamti, L. ; Malach, E. ; Lereclus, D. ; Hayouka, Z. Elucidating the Hot Spot Residues of Quorum Sensing Peptidic Autoinducer PapR by Multiple Amino Acid Replacements. Front Microbiol 2019, 10, 1246.Abstract
The quorum sensing (QS) system of , an opportunistic human pathogen, utilizes the autoinducing PapR peptide signal that mediates the activation of the pleiotropic virulence regulator PlcR. A set of synthetic 7-mer PapR-derived peptides (PapR; ADLPFEF) have been shown to inhibit efficiently the PlcR regulon activity and the production of virulence factors, reflected by a loss in hemolytic activity without affecting bacterial growth. Interestingly, these first potent synthetic inhibitors involved D-amino acid or alanine replacements of three amino acids; proline, glutamic acid, and phenylalanine of the heptapeptide PapR. To better understand the role of these three positions in PlcR activity, we report herein the second generation design, synthesis, and characterization of PapR-derived combinations, alternate double and triple alanine and D-amino acids replacement at these positions. Our findings generate a new set of non-native PapR-derived peptides that inhibit the PlcR regulon activity and the production of virulence factors. Using the amino acids substitution strategy, we revealed the role of proline and glutamic acid on PlcR regulon activation. Moreover, we demonstrated that the D-Glutamic acid substitution was crucial for the design of stronger PlcR antagonists. These peptides represent potent synthetic inhibitors of QS and constitute new and readily accessible chemical tools for the study of the PlcR system. Our method might be applied to other quorum sensing systems to design new anti-virulence agents.