Elongation Factor 1A Family Regulates the Recycling of the M4 Muscarinic Acetylcholine Receptor

In this study, we tested the hypothesis that the elongation 1A (eEF1A) family regulates the cell surface density of the M4 subtype of the muscarinic acetylcholine receptors (mAChR) following agonist-induced internalization. Here, we show that mouse brains lacking eEF1A2 have no detectable changes in M4 expression or localization. We, however, did discover that eEF1A1, the other eEF1A isoform, is expressed in adult neurons contrary to previous reports. This novel finding suggested that the lack of change in M4 expression and distribution in brains lacking eEF1A2 might be due to compensatory effects of eEF1A1. Supporting this theory, we demonstrate that the overexpression of either eEF1A1 or eEF1A2 inhibits M4 recovery to the cell surface after agonist-induced internalization in PC12 cells. Furthermore, eEF1A1 or eEF1A2 had no effect on the recovery of the M1 subtype in PC12 cells. These results demonstrate the novel ability of the eEF1A family to specifically regulate the M4 mAChR.     

Rap1 up-regulation and activation on plasma membrane regulates T cell adhesion

Rap1 and Ras are closely related GTPases that share some effectors but have distinct functions. We studied the subcellular localization of Rap1 and its sites of activation in living cells. Both GFP-tagged Rap1 and endogenous Rap1 were localized to the plasma membrane (PM) and endosomes. The PM association of GFP-Rap1 was dependent on GTP binding, and GFP-Rap1 was rapidly up-regulated on this compartment in response to mitogens, a process blocked by inhibitors of endosome recycling. A novel fluorescent probe for GTP-bound Rap1 revealed that this GTPase was transiently activated only on the PM of both fibroblasts and T cells. Activation on the PM was blocked by inhibitors of endosome recycling. Moreover, inhibition of endosome recycling blocked the ability of Rap1 to promote integrin-mediated adhesion of T cells. Thus, unlike Ras, the membrane localizations of Rap1 are dynamically regulated, and the PM is the principle platform from which Rap1 signaling emanates. These observations may explain some of the biological differences between these GTPases.     

建设资源节约型和环境友好型广东的思考

资源消费量急剧增加,环境压力越来越大,是我国经济社会进一步发展亟待解决的问题。从国家建设资源节约型、环境友好型社会的基本要求出发,结合广东省省情,论述了加快建设资源节约型和环境友好型社会,发展循环经济和促进经济增长方式根本性转变的重要性,认为这是实现广东省经济和社会持续发展,率先基本实现社会主义现代化的必由之路;提出了普及生态科学知识,共同营造生态文明的未来,是建设资源节约型和环境友好型社会的基础。     

Molecular characterization of proprotein convertase subtilisin/kexin type 9-mediated degradation of the LDLR

Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a secreted protein that promotes degradation of cell surface LDL receptors (LDLRs) in selected cell types. Here we used genetic and pharmacological inhibitors to define the pathways involved in PCSK9-mediated LDLR degradation. Inactivating mutations in autosomal recessive hypercholesterolemia (ARH), an endocytic adaptor, blocked PCSK9-mediated LDLR degradation in lymphocytes but not in fibroblasts. Thus, ARH is not specifically required for PCSK9-mediated LDLR degradation. Knockdown of clathrin heavy chain with siRNAs prevented LDLR degradation. In contrast, prevention of ubiquitination of the LDLR cytoplasmic tail, inhibition of proteasomal activity, or disruption of proteins required for lysosomal targeting via macroautophagy (autophagy related 5 and 7) or the endosomal sorting complex required for trafficking (ESCRT) pathway (hepatocyte growth factor-regulated Tyr-kinase substrate and tumor suppressor gene 101) failed to block PCSK9-mediated LDLR degradation. These findings are consistent with a model in which the LDLR-PCSK9 complex is internalized via clathrin-mediated endocytosis and then routed to lysosomes via a mechanism that does not require ubiquitination and is distinct from the autophagy and proteosomal degradation pathways. Finally, the PCSK9-LDLR complex appears not to be transported by the canonical ESCRT pathway.     

A novel recycling process using the treated citric acid wastewater as ingredients water for citric acid production

In this study, an integrated process coupling citric acid and methane fermentations was proposed to solve severe wastewater pollution problem in cassava-based citric acid production. The accumulation patterns of the potential and major inhibitors in this process, including organic compounds, volatile fatty acids (VFAs), total ions and pigments were investigated. Both simulation and experimental results indicated that these inhibitors could reach their equilibrium levels after 3–7 fermentation runs when reutilizing the treated citric acid wastewater. As a result, the proposed citric acid fermentation process by recycling the wastewater treated in methane fermentation could be stably operated for more than 15 runs, which could save a large amount of fresh water and relieve the severe wastewater pollution in citric acid production potentially.     

Critical evaluation of municipal solid waste composting and potential compost markets

Mechanical biological treatment (MBT) of mixed waste streams is becoming increasingly popular as a method for treating municipal solid waste (MSW). Whilst this process can separate many recyclates from mixed waste, the resultant organic residue can contain high levels of heavy metals and physical and biological contaminants. This review assesses the potential end uses and sustainable markets for this organic residue. Critical evaluation reveals that the best option for using this organic resource is in land remediation and restoration schemes. For example, application of MSW-derived composts at acidic heavy metal contaminated sites has ameliorated soil pollution with minimal risk. We conclude that although MSW-derived composts are of low value, they still represent a valuable resource particularly for use in post-industrial environments. A holistic view should be taken when regulating the use of such composts, taking into account the specific situation of application and the environmental pitfalls of alternative disposal routes.     

Class IA PI3Ks regulate subcellular and functional dynamics of IDO1

Knowledge of a protein’s spatial dynamics at the subcellular level is key to understanding its function(s), interactions, and associated intracellular events. Indoleamine 2,3‐dioxygenase 1 (IDO1) is a cytosolic enzyme that controls immune responses via tryptophan metabolism, mainly through its enzymic activity. When phosphorylated, however, IDO1 acts as a signaling molecule in plasmacytoid dendritic cells (pDCs), thus activating genomic effects, ultimately leading to long‐lasting immunosuppression. Whether the two activities—namely, the catalytic and signaling functions—are spatially segregated has been unclear. We found that, under conditions favoring signaling rather than catabolic events, IDO1 shifts from the cytosol to early endosomes. The event requires interaction with class IA phosphoinositide 3‐kinases (PI3Ks), which become activated, resulting in full expression of the immunoregulatory phenotype in vivo in pDCs as resulting from IDO1‐dependent signaling events. Thus, IDO1’s spatial dynamics meet the needs for short‐acting as well as durable mechanisms of immune suppression, both under acute and chronic inflammatory conditions. These data expand the theoretical basis for an IDO1‐centered therapy in inflammation and autoimmunity.     

Scavenger receptor class B type I localizes to a late endosomal compartment

Scavenger receptor class B type I (SR-BI) has an established role in mediating the selective uptake of cholesterol from HDL in hepatocytes, steroidogenic cells, and other tissues. SR-BI is present on the plasma membrane but also localizes to stable intracellular compartments of unknown function. Using indirect immunofluorescence and subcellular fractionation, we have investigated the subcellular distribution of SR-BI. We report that red fluorescent protein-tagged mouse SR-BI (RFP-mSR-BI) colocalizes with the late endosomal and lysosomal markers, Rab7, LBPA, and Rab9. In addition, endogenous SR-BI is also found on lysosomes and colocalizes with LAMP-2 in primary hepatocytes. Furthermore, we demonstrate that the trafficking of SR-BI through these compartments is Rab7 dependent. Interestingly, filipin staining indicates accumulation of lysosomal cholesterol in SR-BI-deficient ((-/-)) as compared with wild-type hepatocytes. In addition to its role as a plasma membrane receptor, SR-BI may function in cholesterol trafficking from late endosomes/lysosomes.     

RhoB-dependent modulation of postendocytic traffic in polarized Madin-Darby canine kidney cells

The Rho family of GTPases is implicated in the control of endocytic and biosynthetic traffic of many cell types; however, the cellular distribution of RhoB remains controversial and its function is not well understood. Using confocal microscopy, we found that endogenous RhoB and green fluorescent protein-tagged wild-type RhoB were localized to early endosomes, and to a much lesser extent to recycling endosomes, late endosomes or Golgi complex of fixed or live polarized Madin-Darby canine kidney cells. Consistent with RhoB localization to early endosomes, we observed that expression of dominant-negative RhoBN19 or dominant-active RhoBV14 altered postendocytic traffic of ligand-receptor complexes that undergo recycling, degradation or transcytosis. In vitro assays established that RhoB modulated the basolateral-to-apical transcytotic pathway by regulating cargo exit from basolateral early endosomes. Our results indicate that RhoB is localized, in part, to early endosomes where it regulates receptor egress through the early endocytic system.