Bipartite binding and partial inhibition links DEPTOR and mTOR in a mutually antagonistic embrace

The mTORC1 kinase complex regulates cell growth, proliferation, and survival. Because mis-regulation of DEPTOR, an endogenous mTORC1 inhibitor, is associated with some cancers, we reconstituted mTORC1 with DEPTOR to understand its function. We find that DEPTOR is a unique partial mTORC1 inhibitor that may have evolved to preserve feedback inhibition of PI3K. Counterintuitively, mTORC1 activated by RHEB or oncogenic mutation is much more potently inhibited by DEPTOR. Although DEPTOR partially inhibits mTORC1, mTORC1 prevents this inhibition by phosphorylating DEPTOR, a mutual antagonism that requires no exogenous factors. Structural analyses of the mTORC1/DEPTOR complex showed DEPTOR’s PDZ domain interacting with the mTOR FAT region, and the unstructured linker preceding the PDZ binding to the mTOR FRB domain. The linker and PDZ form the minimal inhibitory unit, but the N-terminal tandem DEP domains also significantly contribute to inhibition.

Comparative vulnerability of PET radioligands to partial inhibition of P-glycoprotein at the blood-brain barrier: A criterion of choice?

Only partial deficiency/inhibition of P-glycoprotein (P-gp, ABCB1) function at the blood-brain barrier (BBB) is likely to occur in pathophysiological situations or drug-drug interactions. This raises questions regarding the sensitivity of available PET imaging probes to detect moderate changes in P-gp function at the living BBB. In vitro, the half-maximum inhibitory concentration (IC50) of the potent P-gp inhibitor tariquidar in P-gp-overexpressing cells was significantly different using either [11C]verapamil (44 nM), [11C] N-desmethyl-loperamide (19 nM) or [11C]metoclopramide (4 nM) as substrate probes. In vivo PET imaging in rats showed that the half-maximum inhibition of P-gp-mediated efflux of [11C]metoclopramide, achieved using 1 mg/kg tariquidar ( in vivo IC50 = 82 nM in plasma), increased brain exposure by 2.1-fold for [11C]metoclopramide (p < 0.05, n = 4) and 2.4-fold for [11C]verapamil (p < 0.05, n = 4), whereby cerebral uptake of the “avid” substrate [11C] N-desmethyl-loperamide was unaffected (p > 0.05, n = 4). This comparative study points to differences in the “vulnerability” to P-gp inhibition among radiolabeled substrates, which were apparently unrelated to their “avidity” (maximal response to P-gp inhibition). Herein, we advocate that partial inhibition of transporter function, in addition to complete inhibition, should be a primary criterion of evaluation regarding the sensitivity of radiolabeled substrates to detect moderate but physiologically-relevant changes in transporter function in vivo.

Bipartite binding and partial inhibition links DEPTOR and mTOR in a mutually antagonistic embrace

mTORC1 is a kinase complex regulating cell growth, proliferation and survival. Because mis-regulation of DEPTOR, an endogenous mTORC1 inhibitor, is associated with some cancers, we reconstituted mTORC1 with DEPTOR to understand its function. We find that DEPTOR is a unique partial mTORC1 inhibitor that may have evolved to preserve feedback inhibition of PI3K. Counterintuitively, mTORC1 activated by RHEB or oncogenic mutation is much more potently inhibited by DEPTOR. Although DEPTOR partially inhibits mTORC1, mTORC1 prevents this inhibition by phosphorylating DEPTOR, a mutual antagonism that requires no exogenous factors. Structural analyses of the mTORC1/DEPTOR complex showed DEPTOR's PDZ domain interacting with the mTOR FAT region, and the unstructured linker preceding the PDZ binding to the mTOR FRB domain. Here we show, in contrast to previous cellular studies, that both the PDZ and linker regions are essential for inhibition, and it is likely that interaction with the FRB is crucial to the unique partial inhibition.

Identification of the major fermentation inhibitors of recombinant 2G yeasts in diverse lignocellulose hydrolysates

Background Presence of inhibitory chemicals in lignocellulose hydrolysates is a major hurdle for production of second-generation bioethanol. Especially cheaper pre-treatment methods that ensure an economical viable production process generate high levels of these inhibitory chemicals. The effect of several of these inhibitors has been extensively studied with non-xylose-fermenting laboratory strains, in synthetic media, and usually as single inhibitors, or with inhibitor concentrations much higher than those found in lignocellulose hydrolysates. However, the relevance of individual inhibitors in inhibitor-rich lignocellulose hydrolysates has remained unclear. Results The relative importance for inhibition of ethanol fermentation by two industrial second-generation yeast strains in five lignocellulose hydrolysates, from bagasse, corn cobs and spruce, has now been investigated by spiking higher concentrations of each compound in a concentration range relevant for industrial hydrolysates. The strongest inhibition was observed with industrially relevant concentrations of furfural causing partial inhibition of both D-glucose and D-xylose consumption. Addition of 3 or 6 g/L furfural strongly reduced the ethanol titer obtained with strain MD4 in all hydrolysates evaluated, in a range of 34 to 51% and of 77 to 86%, respectively. This was followed by 5-hydroxymethylfurfural, acetic acid and formic acid, for which in general, industrially relevant concentrations caused partial inhibition of D-xylose fermentation. On the other hand, spiking with levulinic acid, 4-hydroxybenzaldehyde, 4-hydroxybenzoic acid or vanillin caused little inhibition compared to unspiked hydrolysate. The further evolved MD4 strain generally showed superior performance compared to the previously developed strain GSE16-T18. Conclusion The results highlight the importance of individual inhibitor evaluation in a medium containing a genuine mix of inhibitors as well as the ethanol that is produced by the fermentation. They also highlight the potential of increasing yeast inhibitor tolerance for improving industrial process economics.

Avena fatua caryopsis dormancy release is associated with changes in KAR1 and ABA sensitivity as well as with ABA reduction in coleorhiza and radicle

Main conclusion The dormancy release in Avena fatua caryopses was associated with a reduction in the ABA content in embryos, coleorhiza and radicle. The coleorhiza proved more sensitive to KAR1 and less sensitive to ABA than the radicle. The inability of dormant caryopses and ABA-treated non-dormant caryopses to complete germination is related to inhibition and delayed of cell-cycle activation, respectively. Abstract As freshly harvested Avena fatua caryopses are dormant at 20 °C, they cannot complete germination; the radicle is not able to emerge. Both karrikin 1 (KAR1) and dry after-ripening release dormancy, enabling the emergence of, first, the coleorhiza and later the radicle. The after-ripening removes caryopse sensitivity to KAR1 and decreases the sensitivity to abscisic acid (ABA). The coleorhiza was found to be more sensitive to KAR1, and less sensitive to ABA, than radicles. Effects of KAR1 and after-ripening were associated with a reduction of the embryo’s ABA content during caryopsis germination. KAR1 was found to decrease the ABA content in the coleorhiza and radicles. Germination of after-ripened caryopses was associated with the progress of cell-cycle activation before coleorhiza emergence. Inhibition of the germination completion due to dormancy or treating the non-dormant caryopses with ABA was associated with a total and partial inhibition of cell-cycle activation, respectively.

Partial inhibition of mitochondrial complex I ameliorates Alzheimer’s disease pathology and cognition in APP/PS1 female mice

Alzheimer’s Disease (AD) is a devastating neurodegenerative disorder without a cure. Here we show that mitochondrial respiratory chain complex I is an important small molecule druggable target in AD. Partial inhibition of complex I triggers the AMP-activated protein kinase-dependent signaling network leading to neuroprotection in symptomatic APP/PS1 female mice, a translational model of AD. Treatment of symptomatic APP/PS1 mice with complex I inhibitor improved energy homeostasis, synaptic activity, long-term potentiation, dendritic spine maturation, cognitive function and proteostasis, and reduced oxidative stress and inflammation in brain and periphery, ultimately blocking the ongoing neurodegeneration. Therapeutic efficacy in vivo was monitored using translational biomarkers FDG-PET, 31P NMR, and metabolomics. Cross-validation of the mouse and the human transcriptomic data from the NIH Accelerating Medicines Partnership–AD database demonstrated that pathways improved by the treatment in APP/PS1 mice, including the immune system response and neurotransmission, represent mechanisms essential for therapeutic efficacy in AD patients.

EFFECT OF EUGENOL EMULSION IN POLYSORBATE-80 ON CLINICAL STRAINS OF CANDIDA ALBICANS

Candida albicans occupies a dominant position in the list of causative agents of candidal lesions of the ear. The development of new antifungal agents, an alternative source of which can be herbal essential oils and their components, remains a priority. One such agent with antiseptic, anti-inflammatory, and analgesic action is eugenol which is a phenol substance. Therefore, this article was aimed to study the effectiveness of the antifungal action of eugenol emulsified in Polysorbate-80 against clinical strains of C. albicans isolated from patients with external otomycosis. The study was performed using 6 clinical strains of C. albicans isolated from the ear canal of patients with otitis externa. The diagnosis of fungal disease was established based on the results of clinical and laboratory (microscopical and mycological) studies of pathological material. Analysis of mycological research showed that mainly representatives of the genus Aspergillus and Penicillium were revealed and only in 7% there were Candida genus fungi. C. albicans remained the dominant species of clinical significance. The results of our studies showed a high level of antifungal activity of eugenol on all clinical strains of C. albicans, including a remarkable inhibitory and fungicidal effect. At postmycostatic concentrations, the eugenol caused partial inhibition of reproduction of the clinical strains of fungi, which was replaced by a subsequent increased cell reproduction rate. So, the investigation has shown that C. albicans is the dominant species among fungi of the Candida genus in the structure of the microbial profile of otomycoses. The eugenol, emulsified in Polysorbate-80, has a high antifungal effect against clinical strains of C. albicans. At postmycostatic concentrations, the eugenol caused partial inhibition of reproduction of the clinical strains of fungi, which was replaced by a subsequent increase cell reproduction rate.

Partial inhibition of borohydride hydrolysis using porous activated carbon as an effective method to improve the electrocatalytic activity of the DBFC anode

Carbon materials are commonly used catalyst supports in various types of fuel cells. Due to the possibility of designing their properties, they seem to be attractive and functional additives. In...