DEPTOR is an inhibitor of the mTOR kinase which controls cell growth. DEPTOR consists of two DEP domains and a PDZ domain connected by an unstructured linker, and its stability is tightly regulated through post-translational modifications of its linker region that contains the 286SSGYFS291 degron. Based on the mTORC1 complex, our modelling suggests a possible spatial arrangement of DEPTOR which is characterised to form a dimer. Our model shows that the two PDZ domains of a DEPTOR dimer bind separately to the dimeric mTOR's FAT domains ~130 Å apart, while each of the two extended linkers is sufficiently long to span from the FAT domain to the kinase domain of mTOR and beyond to join a shared dimer of the DEP domains. This places the linker's S299 closest to the kinase's catalytic site, indicating that phosphorylation would start with it and successively upstream towards DEPTOR's degron. The CK1α kinase is reportedly responsible for the phosphorylation of the degron, and our docking analysis further reveals that CK1α contains sites to bind DEPTOR's pS286, pS287 and pT295, which may act as priming phosphates for the phosphorylation of the degron's S291. DEPTOR's linker can also be ubiquitylated by the UbcH5A-SCFβ-TrCP complex without its PDZ dissociating from mTOR according to the modelling. As the catalytic cleft of mTOR's kinase is restricted, interactions between the kinase's unstructured segment surrounding the cleft and DEPTOR's linker, which may involve S293 and S299, may be critical to controlling DEPTOR's access to the catalytic cleft and hence its phosphorylation by mTOR in a manner dependent on mTOR's activation.
Alzheimer's disease (AD) is a progressive neurological disorder that impairs memory and cognitive abilities, primarily in the elderly. The burden of AD extends beyond patients, impacting families and caregivers due to the patients' reliance on assistance for daily tasks. The main features of the pathogenesis of AD are beta-amyloid plaques and neurofibrillary tangles (NFTs), that strongly correlate with oxidative stress and inflammation. NFTs result from misfolded and hyperphosphorylated tau proteins. Various studies have focused on tau phosphorylation, indicating protein phosphatase 2A (PP2A) as the primary tau phosphatase and glycogen synthase kinase-3 beta (GSK-3β) as the leading tau kinase. Experimental evidence suggests that inhibition of PP2A and increased GSK-3β activity contribute to neuroinflammation, oxidative stress, and cognitive impairment. Hence, targeting PP2A and GSK-3β with pharmacological approaches shows promise in treating AD. The use of natural compounds in the drug development for AD have been extensively studied for their antioxidant, anti-inflammatory, anti-cholinesterase, and neuroprotective properties, demonstrating therapeutic advantages in neurological diseases. Alongside the development of PP2A activator and GSK-3β inhibitor drugs, natural compounds are likely to have neuroprotective effects by increasing PP2A activity and decreasing GSK-3β levels. Therefore, based on the preclinical and clinical studies, the potential of PP2A and GSK-3β as therapeutic targets of natural compounds are highlighted in this review.
Endothelial barrier dysfunction leads to increased endothelial permeability and is an early step in the development of vascular inflammatory diseases such as atherosclerosis. Interferon-γ (IFN-γ), a proinflammatory cytokine, is known to cause increased endothelial permeability. However, the mechanisms by which IFN-γ disrupts the endothelial barrier have not been clarified. This study aimed to investigate how IFN-γ impairs the endothelial barrier integrity by specifically examining the roles of caldesmon, adherens junctions (AJs) and p38 mitogen-activated protein (MAP) kinase in IFN-γ-induced endothelial barrier dysfunction. IFN-γ exhibited a biphasic effect on caldesmon localization and both the structural organization and protein expression of AJs. In the early phase (4-8 h), IFN-γ induced the formation of peripheral caldesmon bands and discontinuous AJs, while AJ protein expression was unchanged. Interestingly, IFN-γ also stimulated caldesmon phosphorylation, resulting in actin dissociation from caldesmon at 8 h. Conversely, changes seen in the late phase (16-24 h) included cytoplasmic caldesmon dispersal, AJ linearization and junctional area reduction, which were associated with reduced membrane, cytoskeletal and total AJ protein expression. In addition, IFN-γ enhanced myosin binding to caldesmon at 12 h and persisted up to 24 h. Furthermore, inhibition of p38 MAP kinase by SB203580 did not reverse either the early or late phase changes observed. These data suggest that IFN-γ may activate signaling molecules other than p38 MAP kinase. In conclusion, our findings enhance the current understanding of how IFN-γ disrupts endothelial barrier function and reveal potential therapeutic targets, such as caldesmon and AJs, for the treatment of IFN-γ-associated vascular inflammatory diseases.
This prospective observational study was to evaluate the efficacy of combining phosphorylated insulin-like growth factor binding protein-1 (phIGFBP-1) and transvaginal ultrasound cervical length (CL) compared with either indicator alone in predicting pre-term labour (PTL). Women with singleton pregnancy between 24 and 36 weeks' gestation with evidence of PTL were subjected to phIGFBP-1 and CL tests. Of the 51 women, five were tested positive (phIGFBP-1 positive and CL <2.5 cm) for combination of phIGFBP-1 and CL (four delivered within 1 week), whereas 46 tested negative, of which, only one delivered. A much higher negative predictivity (NP), positive predictivity (PP) and specificity (SP) in the combination test was seen compared with phIGFBP-1 or CL alone (NP: 97.8% vs 97.7% vs 97.1%; PP: 80.0% vs 51.1% and CL 23.5%; SP: 97.8% vs 93.5% vs 71.1%, respectively). The cervical os dilatation of 2 cm with combined positive test (p = 0.001) indicated a higher likelihood of PTL.
We determined the effects of "periodized nutrition" on skeletal muscle and whole body responses to a bout of prolonged exercise the following morning. Seven cyclists completed two trials receiving isoenergetic diets differing in the timing of ingestion: they consumed either 8 g/kg body mass (BM) of carbohydrate (CHO) before undertaking an evening session of high-intensity training (HIT) and slept without eating (FASTED), or consumed 4 g/kg BM of CHO before HIT, then 4 g/kg BM of CHO before sleeping (FED). The next morning subjects completed 2 h of cycling (120SS) while overnight fasted. Muscle biopsies were taken on day 1 (D1) before and 2 h after HIT and on day 2 (D2) pre-, post-, and 4 h after 120SS. Muscle [glycogen] was higher in FED at all times post-HIT (P < 0.001). The cycling bouts increased PGC1α mRNA and PDK4 mRNA (P < 0.01) in both trials, with PDK4 mRNA being elevated to a greater extent in FASTED (P < 0.05). Resting phosphorylation of AMPK(Thr172), p38MAPK(Thr180/Tyr182), and p-ACC(Ser79) (D2) was greater in FASTED (P < 0.05). Fat oxidation during 120SS was higher in FASTED (P = 0.01), coinciding with increases in ACC(Ser79) and CPT1 as well as mRNA expression of CD36 and FABP3 (P < 0.05). Methylation on the gene promoter for COX4I1 and FABP3 increased 4 h after 120SS in both trials, whereas methylation of the PPARδ promoter increased only in FASTED. We provide evidence for shifts in DNA methylation that correspond with inverse changes in transcription for metabolically adaptive genes, although delaying postexercise feeding failed to augment markers of mitochondrial biogenesis.
The cognitive impairment caused by Alzheimer's disease (AD) is associated with beta-amyloid (Aβ) and tau proteins, and is accompanied by inflammation. Recently, a novel inflammasome signaling pathway has been uncovered. Inflammasomes are implicated in the execution of inflammatory responses and pyroptotic death leading to neurodegeneration. Thus, the inflammasome signaling pathway could be a potential therapeutic target for AD. Neural stem cells (NSCs) are multipotent cells that can self-renew and differentiate into distinct neural cells. NSC therapy has been considered to be a promising therapeutic approach in protecting the central nervous system and restoring it following damage. However, the mechanisms involved remain unclear. The aims of this study were to investigate the protective effects of NE4C neural stem cells against microglia-mediated neurotoxicity and to explore molecular mechanisms mediating their actions. NE4C decreased the levels of caspase-1 and IL-1β, and attenuated the level of the NLRP3 inflammasome and its associated protein adapter, apoptosis-associated speck-like protein containing a C-terminal caspase recruitment domain (ASC) in LPS-stimulated BV2 microglial cells, possibly by regulating the phosphorylation of p38α MAPK. The conditioned media obtained from co-culture of LPS-stimulated BV2 and NE4C cells exhibited protective effects on SH-SY5Y cells against microglia-mediated neurotoxicity; this was associated with an attenuation of tau phosphorylation and amyloidogenesis and accompanied by down-regulation of GSK-3β and p38α MAPK signalling pathways. In conclusion, the present study suggested that NSC therapy could be a potential strategy against microglia-mediated neurotoxicity. NSCs regulate NLRP3 activation and IL-1β secretion, which are critical in the initiation of the inflammatory responses, hence preventing the release of neurotoxic pro-inflammatory factors by microglia. This eventually reduces tau hyperphosphylation and amyloidogenesis, possibly through the regulation of GSK-3β and p38α MAPK signalling pathways, and thus protects SH-SY5Y cells against microglia-mediated neurotoxicity.