Super-resolution optical microscopy has extended the spatial resolution of cell biology from the cellular level to the nanoscale, enabling the observation of the interactive behavior of single ...mitochondria and lysosomes. Quantitative parametrization of interactions between mitochondria and lysosomes under super-resolution optical microscopy, however, is currently unavailable, which has severely limited our understanding of the molecular machinery underlying mitochondrial functionality. Here, we introduce an M-value to quantitatively investigate mitochondria and lysosome contact (MLC) and mitophagy under structured illumination microscopy. We found that the M-value for an MLC is typically less than 0.4, whereas in mitophagy it ranges from 0.5 to 1.0. This system permits further investigation of the detailed molecular mechanism governing the interactive behavior of mitochondria and lysosomes.
FIP200 is an essential autophagy gene implicated in the regulation of postnatal neural progenitor/stem cells (NSCs). However, the contribution of FIP200's canonical-autophagy function and its ...non-canonical functions to postnatal NSC maintenance remains unclear. Utilizing a recently generated Fip200-4A allele that specifically impairs FIP200's canonical-autophagy function, we found that non-canonical functions of FIP200 was required for regulation of mouse NSC maintenance and neurogenesis in vivo. Ablating the non-canonical functions of FIP200, but not its autophagy function, increased TBK1 activation and p62 phosphorylation at S403 in NSCs. Phosphorylation of p62 was dependent on TBK1 kinase activity and increased the propensity of p62 aggregate formation specifically in FIP200-null NSCs. Accordingly, inhibition of TBK1 by amlexanox reduced p62 aggregates and restored NSC maintenance and differentiation in Fip200
cKO mice. These results reveal a mechanism for the non-canonical functions of FIP200 in NSC maintenance and differentiation by limiting TBK1 activation and subsequently, p62 aggregate formation.
Betulinic acid (BA) and its derivatives are a class of high-profile drug candidates, but their anticancer effects on resistant cancer have rarely been reported. Although a few studies indicated ...mitophagy is related with drug resistance, its role in different cancer types and anticancer agents treatment remains largely unclear. Here, we find that B5G1, a new derivative of BA, induces cell death in multidrug resistant cancer cells HepG2/ADM and MCF-7/ADR through mitochondrial-apoptosis pathway. B5G1 also triggers mitophagy independent on Atg5/Beclin 1. Further mechanistic study indicates that B5G1 upregulates PTEN-induced putative kinase 1 (PINK1) to recruit Parkin to mitochondria followed by ubiquitination of Mfn2 to initiate mitophagy. Inhibition of mitophagy by PINK1 siRNA, mdivi-1, or bafilomycin A1 (Baf A1) promotes B5G1-induced cell death. In addition, ROS production and mitochondrial damage in B5G1-treated HepG2/ADM cells cause mitochondrial apoptosis and mitophagy. In vivo study shown that B5G1 dramatically inhibits HepG2/ADM xenograft growth accompanied by apoptosis and mitophagy induction. Together, our results provide the first demonstration that B5G1, as a novel mitophagy inducer, has the potential to be developed into a drug candidate for treating multidrug resistant cancer.
Recent studies have shown important roles for autophagy genes in the regulation of different tissue stem cells, including neural stem/progenitor cells (NSCs). However, little is known about whether ...autophagy can regulate NSCs through cell-extrinsic mechanisms. Here, we show that deletion of an essential autophagy gene, FIP200, in NSCs increased expression of Ccl5 and Cxcl10 in a p53-independent manner, mediating increased infiltration of microglia into the subventricular zone of both FIP200hGFAP conditional knockout (cKO) and FIP200;p53hGFAP 2cKO mice. The microglia exhibited an activated M1 phenotype consistent with their potential to inhibit differentiation of FIP200-null NSCs. Blocking either microglia infiltration or activation rescued the deficient differentiation of FIP200-null NSCs from FIP200;p53hGFAP 2cKO mice. Lastly, we showed that increased chemokine expression in FIP200-null NSCs was induced by abnormal p62 aggregate formation and activation of NF-κB signaling. Our results suggest that autophagy plays a crucial role in regulating neurogenesis and restricting local immune response in postnatal NSCs through non-cell autonomous mechanisms.
Protein ligand docking is an indispensable tool for computational prediction of protein functions and screening drug candidates. Despite significant progress over the past two decades, it is still a ...challenging problem, characterized by the still limited understanding of the energetics between proteins and ligands, and the vast conformational space that has to be searched to find a satisfactory solution. In this project, we developed a novel reinforcement learning (RL) approach, the asynchronous advantage actor-critic model (A3C), to address the protein ligand docking problem. The overall framework consists of two models. During the search process, the agent takes an action selected by the actor model based on the current location. The critic model then evaluates this action and predict the distance between the current location and true binding site. Experimental results showed that in both single- and multi-atom cases, our model improves binding site prediction substantially compared to a naïve model. For the single-atom ligand, copper ion (Cu.sup.2+), the model predicted binding sites have a median root-mean-square-deviation (RMSD) of 2.39 Ã to the true binding sites when starting from random starting locations. For the multi-atom ligand, sulfate ion (SO.sub.4.sup.2-), the predicted binding sites have a median RMSD of 3.82 Ã to the true binding sites. The ligand-specific models built in this study can be used in solvent mapping studies and the RL framework can be readily scaled up to larger and more diverse sets of ligands.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The mechanistic target of rapamycin complex 1 (mTORC1) is important in the development and progression of many cancers. Targeted cancer therapy using mTORC1 inhibitors is used for treatment of ...cancers; however, their clinical efficacies are still limited.
We recently created a new mouse model for human lymphangiosarcoma by deleting Tsc1 in endothelial cells and consequent hyper-activation of mTORC1. Using Tsc1
tumour cells from this mouse model, we assessed the efficacies of histone deacetylase (HDAC) inhibitors as anti-tumour agents for mTORC1-driven tumours.
Unlike the cytostatic effect of mTORC1 inhibitors, HDAC inhibitors induced Tsc1
tumour cell death in vitro and their growth in vivo. Analysis of several HDAC inhibitors suggested stronger anti-tumour activity of class I HDAC inhibitor than class IIa or class IIb inhibitors, but these or pan HDAC inhibitor SAHA did not affect mTORC1 activation in these cells. Moreover, HDAC inhibitor-induced cell death required elevated autophagy, but was not affected by disrupting caspase-dependent apoptosis pathways. We also observed increased reactive oxygen species and endoplasmic reticulum stress in SAHA-treated tumour cells, suggesting their contribution to autophagic cell death, which were dependent on mTORC1 hyper-activation.
These studies suggest a potential new treatment strategy for mTORC1-driven cancers like lymphangiosarcoma through an alternative mechanism.
Background Ischemic heart disease (IHD) imposes the greatest disease burden globally, especially in low- and middle-income countries (LMICs). We aim to examine the population-attributable fraction ...and risk-attributable death and disability-adjusted life years (DALYs) for IHD in 137 low- and middle-income countries. Methods and Results Using comparative risk assessment framework from the 2019 Global Burden of Disease study, the population-attributable fraction and IHD burden (death and DALYs) attributable to risk factors in low-income countries, lower-middle-income countries (LMCs), and upper-middle-income countries were assessed from 2000 to 2019. In 2019, the population-attributable fraction (%) of IHD deaths in relation to all modifiable risk factors combined was highest in lower-middle-income countries (94.2; 95% uncertainty interval, 91.9-96.2), followed by upper-middle-income countries (93.5; 90.4-95.8) and low-income countries (92.5; 90.0-94.7). There was a >13-fold difference between Peru and Uzbekistan in age-standardized rates (per 100 000) of attributable death (44.3 versus 660.4) and DALYs (786.7 versus 10506.1). Dietary risks accounted for the largest proportion of IHD's behavioral burden in low- and middle-income countries, primarily attributable to diets low in whole grains. High systolic blood pressure and high low-density lipoprotein cholesterol remained the 2 leading causes of DALYs, with the former topping the list in 116 countries, while the latter led in 21 of the 137 countries. Compared with 2000 to 2010, the increases in risk-attributable deaths and DALYs among upper-middle income countries were slower from 2010 to 2019, while the trends in low-income countries and lower-middle income countries were opposite. Conclusions IHD's attributable burden remains high in low- and middle-income countries. Considerable heterogeneity was observed among different income-classified regions and countries.
Tuberous sclerosis complex (TSC) is a genetic disorder caused by inactivating mutations in TSC1 (hamartin) or TSC2 (tuberin), crucial negative regulators of the mechanistic target of rapamycin ...complex 1 (mTORC1) signaling pathway. TSC affects multiple organs including the brain. The neurologic manifestation is characterized by cortical tubers, subependymal nodules (SEN), and subependymal giant cell astrocytoma (SEGA) in brain. SEGAs may result in hydrocephalus in TSC patients and mTORC1 inhibitors are the current recommended therapy for SEGA. Nevertheless, a major limitation in the research for SEGA is the lack of cell lines or animal models for mechanistic investigations and development of novel therapy. In this study, we generated TSC1-deficient neural cells from spontaneously immortalized mouse astrocytes in an attempt to mimic human SEGA. The TSC1-deficient cells exhibit mTORC1 hyperactivation and characteristics of transition from astrocytes to neural stem/progenitor cell phenotypes. Rapamycin efficiently decreased mTORC1 activity of these TSC1-deficient cells in vitro. In vivo, TSC1-deficient cells could form SEGA-like tumors and Rapamycin treatment decreased tumor growth. Collectively, our study generates a novel SEGA-like cell line that is invaluable for studying mTORC1-driven molecular and pathological alterations in neurologic tissue. These SEGA-like cells also provide opportunities for the development of novel therapeutic strategy for TSC patients with SEGA.
FIP200 (FAK family-interacting protein of 200 kDa) is a conserved protein recently identified as a potential mammalian counterpart of yeast autophagy protein Atg17. However, it remains unknown ...whether mammalian FIP200 regulates autophagy in vivo. Here we show that neural-specific deletion of FIP200 resulted in cerebellar degeneration accompanied by progressive neuronal loss, spongiosis, and neurite degeneration in the cerebellum. Furthermore, deletion of FIP200 led to increased apoptosis in cerebellum as well as accumulation of ubiquitinated protein aggregates without any deficiency in proteasome catalytic functions. We also observed an increased p62/SQSTM1 accumulation in the cerebellum and reduced autophagosome formation as well as accumulation of damaged mitochondria in the mutant mice. Lastly, analysis of cerebellar neurons in vitro showed reduced JNK activation and increased susceptibility to serum deprivation-induced apoptosis in cerebellar neurons from the mutant mice. Taken together, these results provide strong genetic evidence for a role of FIP200 in the regulation of neuronal homeostasis through its function in autophagy in vivo.
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