Multicomponent Characterization of the Flower Bud of Panax notoginseng and Its Metabolites in Rat Plasma by Ultra-High Performance Liquid Chromatography/Ion Mobility Quadrupole Time-of-Flight Mass Spectrometry

The flower bud of Panax notoginseng (PNF) consumed as a tonic shows potential in the prevention and treatment of cardiovascular diseases. To identify the contained multi-components and, in particular, to clarify which components can be absorbed and what metabolites are transformed, unveiling the effective substances of PNF is of vital significance. A unique ultrahigh-performance liquid chromatography/ion mobility quadrupole time-of-flight mass spectrometry (UHPLC/IM-QTOF-MS) profiling approach and efficient data processing by the UNIFITM bioinformatics platform were employed to comprehensively identify the multi-components of PNF and the related metabolites in the plasma of rats after oral administration (at a dose of 3.6 g/kg). Two MS2 data acquisition modes operating in the negative electrospray ionization mode, involving high-definition MSE (HDMSE) and data-dependent acquisition (DDA), were utilized aimed to extend the coverage and simultaneously ensure the quality of the MS2 spectra. As a result, 219 components from PNF were identified or tentatively characterized, and 40 thereof could be absorbed. Moreover, 11 metabolites were characterized from the rat plasma. The metabolic pathways mainly included the phase I (deglycosylation and oxidation). To the best of our knowledge, this is the first report that systematically studies the in vivo metabolites of PNF, which can assist in better understanding its tonifying effects and benefit its further development

MTORC1-mediated NRBF2 phosphorylation functions as a switch for the class III PtdIns3K and autophagy

NRBF2/Atg38 has been identified as the fifth subunit of the macroautophagic/autophagic class III phosphatidylinositol 3-kinase (PtdIns3K) complex, along with ATG14/Barkor, BECN1/Vps30, PIK3R4/p150/Vps15 and PIK3C3/Vps34. However, its functional mechanism and regulation are not fully understood. Here, we report that NRBF2 is a fine tuning regulator of PtdIns3K controlled by phosphorylation. Human NRBF2 is phosphorylated by MTORC1 at S113 and S120. Upon nutrient starvation or MTORC1 inhibition, NRBF2 phosphorylation is diminished. Phosphorylated NRBF2 preferentially interacts with PIK3C3/PIK3R4. Suppression of NRBF2 phosphorylation by MTORC1 inhibition alters its binding preference from PIK3C3/PIK3R4 to ATG14/BECN1, leading to increased autophagic PtdIns3K complex assembly, as well as enhancement of ULK1 protein complex association. Consequently, NRBF2 in its unphosphorylated form promotes PtdIns3K lipid kinase activity and autophagy flux, whereas its phosphorylated form blocks them. This study reveals NRBF2 as a critical molecular switch of PtdIns3K and autophagy activation, and its on/off state is precisely controlled by MTORC1 through phosphorylation

The exon 19-deleted EGFR undergoes ubiquitylation-mediated endocytic degradation via dynamin activity-dependent and -independent mechanisms

Abstract Background The epidermal growth factor receptor (EGFR) is closely implicated in cancer, and sequencing analyses have revealed a high mutation rate of EGFR in lung cancer. Recent advances have provided novel insights into the endocytic regulation of wild-type EGFR, but that of mutated EGFR remains elusive. In the present study, we aim to investigate the endocytic degradation of a frequently occurred exon 19-deleted mutant in lung cancer. Methods The EGF-induced endocytic degradation of EGFR was examined in a panel of lung cancer cells using immunoblotting. The subcellular distribution of internalized EGFR was investigated using immunofluorescence and confocal microscopy. The effects of dynamin were assessed using its small molecule inhibitors, while the influence of RTN3 was tested using shRNA-mediated knockdown. Finally the ubiquitylation status of EGFR mutant was studied using immunoprecipitation under steady state and tyrosine kinase inhibitor-treated conditions. Results EGF induced various rates of EGFR endocytic degradation in lung cancer cells. Interestingly, the exon 19 deletion mutant is constantly internalized and sorted to lysosome for degradation, and this process is independent of dynamin activity. EGF stimulation and HSP90 inhibition further enhance the endocytic degradation of the exon 19 deletion mutant, in a dynamin activity-dependent and -independent manner, respectively. Albeit with different modes of internalization, the uptake of the exon 19-deleted EGFR is mediated through receptor ubiquitylation. Conclusions The internalized EGFR mutant is constantly routed through endosome to lysosome for degradation. The endocytosis of EGFR mutant occurs through both dynamin activity-dependent and -independent mechanisms. Our findings gain novel insights into the endocytic regulation of mutated EGFR and may have potential clinical implications

Integrated opposite charge grafting induced ionic-junction fiber

The emergence of ionic-junction devices has attracted growing interests due to the potential of serving as signal transmission and translation media between electronic devices and biological systems using ions. Among them, fiber-shaped iontronics possesses a great advantage in implantable applications owing to the unique one-dimensional geometry. However, fabricating stable ionic-junction on curved surfaces remains a challenge. Here, we developed a polyelectrolyte based ionic-junction fiber via an integrated opposite charge grafting method capable of large-scale continuous fabrication. The ionic-junction fibers can be integrated into functions such as ionic diodes and ionic bipolar junction transistors, where rectification and switching of input signals are implemented. Moreover, synaptic functionality has also been demonstrated by utilizing the fiber memory capacitance. The connection between the ionic-junction fiber and sciatic nerves of the mouse simulating end-to-side anastomosis is further performed to realize effective nerve signal conduction, verifying the capability for next-generation artificial neural pathways in implantable bioelectronics. Ionic-junction devices are difficult to integrate with fiber-shaped tissues like nerves and muscles for applications in implantable bioelectronics due to their large size and bulk structure. Authors realize here easy to implant fiber-shaped iontronics through an integrated opposite charge grafting process, enabling the construction of ionic logic gates and artificial neural pathways.Funding Agencies|National Natural Science Foundation of China [92163132]; Fundamental Research Funds for the Central Universities [2232020A-03, 2232021G-12]; NSFC [52173156]; Science and Technology Commission of Shanghai Municipality [20JC1414900, 19JC1410600, 21520710700]; National Key Laboratory Foundation of China [JCKYS2022LD7]; VINNOVA [2020-05223]; Fundamental Research Funds for the Central Universities and Graduate Student Innovation Fund of Donghua University [CUSF-DH-D-2022002]</p