mTORC1 activity is supported by spatial association with focal adhesions

The mammalian target of rapamycin complex 1 (mTORC1) integrates mitogenic and stress signals to control growth and metabolism. Activation of mTORC1 by amino acids and growth factors involves recruitment of the complex to the lysosomal membrane and is further supported by lysosome distribution to the cell periphery. Here, we show that translocation of lysosomes toward the cell periphery brings mTORC1 into proximity with focal adhesions (FAs). We demonstrate that FAs constitute discrete plasma membrane hubs mediating growth factor signaling and amino acid input into the cell. FAs, as well as the translocation of lysosome-bound mTORC1 to their vicinity, contribute to both peripheral and intracellular mTORC1 activity. Conversely, lysosomal distribution to the cell periphery is dispensable for the activation of mTORC1 constitutively targeted to FAs. This study advances our understanding of spatial mTORC1 regulation by demonstrating that the localization of mTORC1 to FAs is both necessary and sufficient for its activation by growth-promoting stimuli.     

The cephalic morphology of the Gondwanan key taxon Hackeriella (Coleorrhyncha,Hemiptera)

External and internal head structures of Coleorrhyncha, a key-taxon within the Hemiptera, are described in detail and documented using modern techniques. The main focus is on Hackeriella veitchi, but two additional representatives of the Gondwanan relict group were also examined, and also head structures of Enicocephalidae, a member of a potentially basal heteropteran lineage. Features were compared to those documented in literature for the Sternorrhyncha, Auchenorrhyncha, and Heteroptera. Coleorrhyncha are characterized by highly modified head structures and correspondingly an entire series of autapomorphies, such as for instance a strongly flattened head capsule with fenestrations. However, they also display features that are likely plesiomorphic compared to members of other hemipteran groups. These include the almost complete tentorium and the lack of the gula. The sistergroup relationship between Coleorrhyncha and Heteroptera is well supported by cephalic features. Potential synapomorphies are the presence of a distinct mandibular sulcus, the reduced number of antennomeres, the absence of clasping organs in the labial groove, coiled accessory salivary ducts, the presence of a small cervical muscle M1a (M. pronotopostoccipitalis medialis), the presence of a second mandibular promotor M14 (M. zygomaticus mandibulae), the presence of M28 (M. verticopharyngalis), and M30 (M. frontobuccalis posterior).     

Accelerated Activation of SOCE Current in Myotubes from Two Mouse Models of Anesthetic- and Heat-Induced Sudden Death

Store-operated calcium entry (SOCE) channels play an important role in Ca²⁺ signaling. Recently, excessive SOCE was proposed to play a central role in the pathogenesis of malignant hyperthermia (MH), a pharmacogenic disorder of skeletal muscle. We tested this hypothesis by characterizing SOCE current (I_SkCRAC) magnitude, voltage dependence, and rate of activation in myotubes derived from two mouse models of anesthetic- and heat-induced sudden death: 1) type 1 ryanodine receptor (RyR1) knock-in mice (Y524S/+) and 2) calsequestrin 1 and 2 double knock-out (dCasq-null) mice. I_SkCRAC voltage dependence and magnitude at -80 mV were not significantly different in myotubes derived from wild type (WT), Y524S/+ and dCasq-null mice. However, the rate of I_SkCRAC activation upon repetitive depolarization was significantly faster at room temperature in myotubes from Y524S/+ and dCasq-null mice. In addition, the maximum rate of I_SkCRAC activation in dCasq-null myotubes was also faster than WT at more physiological temperatures (35-37°C). Azumolene (50 µM), a more water-soluble analog of dantrolene that is used to reverse MH crises, failed to alter I_SkCRAC density or rate of activation. Together, these results indicate that while an increased rate of I_SkCRAC activation is a common characteristic of myotubes derived from Y524S/+ and dCasq-null mice and that the protective effects of azumolene are not due to a direct inhibition of SOCE channels.     

Membrane protein shaving with thermolysin can be used to evaluate topology predictors

Topology analysis of membrane proteins can be obtained by enzymatic shaving in combination with MS identification of peptides. Ideally, such analysis could provide quite detailed information about the membrane spanning regions. Here, we examine the ability of some shaving enzymes to provide large‐scale analysis of membrane proteome topologies. To compare different shaving enzymes, we first analyzed the detected peptides from two over‐expressed proteins. Second, we analyzed the peptides from non‐over‐expressed Escherichia coli membrane proteins with known structure to evaluate the shaving methods. Finally, the identified peptides were used to test the accuracy of a number of topology predictors. At the end we suggest that the usage of thermolysin, an enzyme working at the natural pH of the cell for membrane shaving, is superior because: (i) we detect a similar number of peptides and proteins using thermolysin and trypsin; (ii) thermolysin shaving can be run at a natural pH and (iii) the incubation time is quite short. (iv) Fewer detected peptides from thermolysin shaving originate from the transmembrane regions. Using thermolysin shaving we can also provide a clear separation between the best and the less accurate topology predictors, indicating that using data from shaving can provide valuable information when developing new topology predictors.     

The Natural Product Magnolol as a Lead Structure for the Development of Potent Cannabinoid Receptor Agonists

Magnolol (4-allyl-2-(5-allyl-2-hydroxyphenyl)phenol), the main bioactive constituent of the medicinal plant Magnolia officinalis, and its main metabolite tetrahydromagnolol were recently found to activate cannabinoid (CB) receptors. We now investigated the structure-activity relationships of (tetrahydro)magnolol analogs with variations of the alkyl chains and the phenolic groups and could considerably improve potency. Among the most potent compounds were the dual CB₁/CB₂ full agonist 2-(2-methoxy-5-propyl-phenyl)-4-hexylphenol (61a, K _i CB₁∶0.00957 µM; K _i CB₂∶0.0238 µM), and the CB₂-selective partial agonist 2-(2-hydroxy-5-propylphenyl)-4-pentylphenol (60, K _i CB₁∶0.362 µM; K _i CB₂∶0.0371 µM), which showed high selectivity versus GPR18 and GPR55. Compound 61b, an isomer of 61a, was the most potent GPR55 antagonist with an IC₅₀ value of 3.25 µM but was non-selective. The relatively simple structures, which possess no stereocenters, are easily accessible in a four- to five-step synthetic procedure from common starting materials. The central reaction step is the well-elaborated Suzuki-Miyaura cross-coupling reaction, which is suitable for a combinatorial chemistry approach. The scaffold is versatile and may be fine-tuned to obtain a broad range of receptor affinities, selectivities and efficacies.     

Pristine C60 Fullerene Nanoparticles Ameliorate Hyperglycemia-Induced Disturbances via Modulation of Apoptosis and Autophagy Flux

Neurochemical Research - Diabetes mellitus is a prevalent metabolic disorder associated with multiple complications including neuropathy, memory loss and cognitive decline. Despite a long history...     

Host genetic risk factors for community-acquired pneumonia

This study was conducted to establish the contribution of genetic host factors in the susceptibility to community acquired pneumonia (CAP) in the Russian population. Patients with CAP (n = 334), volunteers without a previous history of CAP, constantly exposed to infectious agents, control A group (n = 141) and a second control group B consisted of healthy persons (n = 314) were included in the study. All subjects were genotyped for 13 polymorphic variants in the genes of xenobiotics detoxification CYP1A1 (rs2606345, rs4646903, and rs1048943), GSTM1 (Ins/del), GSTT1 (Ins/del), ABCB1 rs1045642); immune and inflammation response IL-6 (rs1800795), TNF-a (rs1800629), MBL2 (rs7096206), CCR5 (rs333), NOS3 (rs1799983), angiotensin-converting enzyme ACE (rs4340), and occlusive vascular disease/hyperhomocysteinemia MTHFR (rs1801133). Seven polymorphic variants in genes CYP1A1, GSTM1, ABCB1, NOS3, IL6, CCR5 and ACE were associated with CAP. For two genes CYP1A1 and GSTM1 associations remained significant after correction for multiple comparisons. Multiple analysis by the number of all risk genotypes showed a highly significant association with CAP (P = 2.4 × 10^− 7, OR = 3.03, 95% CI 1.98–4.64) with the threshold for three risk genotypes. Using the ROC-analysis, the AUC value for multi-locus model was estimated as 68.38.