Tyrosine phosphorylation directs TACE into extracellular vesicles via unconventional secretion

When studying how HIV‐1 Nef can promote packaging of the proinflammatory transmembrane protease TACE (tumor necrosis factor‐α converting enzyme) into extracellular vesicles (EVs) we have revealed a novel tyrosine kinase‐regulated unconventional protein secretion (UPS) pathway for TACE. When TACE was expressed without its trafficking cofactor iRhom allosteric Hck activation by Nef triggered translocation of TACE into EVs. This process was insensitive to blocking of classical secretion by inhibiting endoplasmic reticulum (ER) to Golgi transport, and involved a distinct form of TACE devoid of normal glycosylation and incompletely processed for prodomain removal. Like most other examples of UPS this process was Golgi reassembly stacking protein (GRASP)‐dependent but was not associated with ER stress. These data indicate that Hck‐activated UPS provides an alternative pathway for TACE secretion that can bypass iRhom‐dependent ER to Golgi transfer, and suggest that tyrosine phosphorylation might have a more general role in regulating UPS.      

The overexpression of GRASP might inhibit cell proliferation and invasion in hepatocellular carcinoma

This study aimed to validate the methylation of key genes in hepatocellular carcinoma (HCC) screened by bioinformatics analysis and explore whether they affected HCC cell proliferation, migration, and invasion. Using The Cancer Genome Atlas (TCGA) database, HCC-related differentially methylated positions (DMPs) were screened, genes corresponding to DMPs were selected, and prognosis-related genes were identified. A representative DMP was used to divide the DMPs into hyper- and hypomethylated groups. Expression of key genes in cell lines was detected using quantitative real-time polymerase chain reaction and western blot analysis. After treatment of HepG2 cells with 5-Aza-2′-deoxycytidine (5-Aza-DC), gene expression was observed. Bisulfite sequencing PCR assay was used to detect methylation frequency. Overexpressed GRASP lentiviral vectors were constructed to analyze their influence on cell proliferation, migration, and invasion using cell counting kit-8 and transwell assays. Forty-three HCC prognosis-related genes were screened using the TCGA database. cg00249511 (SCT) was used to divide the DMPs into hyper- and hypomethylated groups, distinguishing between high- and low-risk samples. The prognosis survival model constructed using 12 genes revealed the prognosis type. GRASP messenger RNA was downregulated in HepG2 and upregulated after 5-Aza-DC treatment. In HCC tissues, methylation frequency of GRASP was upregulated. GRASP overexpression inhibited HepG2 cell proliferation, invasion, and G-CSFR expression. Thus, GRASP might be a prognosis-related gene controlled by methylation.     

Golgi duplication in Trypanosoma brucei

Duplication of the single Golgi apparatus in the protozoan parasite Trypanosoma brucei has been followed by tagging a putative Golgi enzyme and a matrix protein with variants of GFP. Video microscopy shows that the new Golgi appears de novo, near to the old Golgi, about two hours into the cell cycle and grows over a two-hour period until it is the same size as the old Golgi. Duplication of the endoplasmic reticulum (ER) export site follows exactly the same time course. Photobleaching experiments show that the new Golgi is not the exclusive product of the new ER export site. Rather, it is supplied, at least in part, by material directly from the old Golgi. Pharmacological experiments show that the site of the new Golgi and ER export is determined by the location of the new basal body.     

Spatially modelling native vegetation condition

Summary   The assessment of vegetation condition is seen as an increasingly important requirement for effective biodiversity conservation in Australia. Condition assessments that operate at the scale of the site are well established. However, there is a need for mapped representations of vegetation condition at regional scales to: (i) assist with regional planning and target setting; (ii) provide regional context for site-based assessment; and (iii) monitor the change in vegetation condition at multiple scales. This paper describes a methodology for converting site condition data collected in plots into maps of vegetation condition across entire regions using a predictive statistical modelling framework (Generalized Additive Modelling) combined with a GIS. The research demonstrates how explanatory variables including topographic position, terrain roughness, landscape connectivity and remote sensing derived indices can be used to map the condition of native vegetation at the scale of a subcatchment. The inclusion of indices derived from remotely sensed imagery (SPOT4) as explanatory variables in the modelling is a novel component of this research. Although the methodology generates statistically and ecologically plausible models of vegetation condition, there are nevertheless limitations associated with the way plot data were collected and some of the explanatory variables, which impacts upon model utility. We discuss how these problems can be minimized when embarking upon studies of this type. We demonstrate how maps produced from exercises such as this could be used for conservation planning and discuss the limitations of these data for monitoring.     

Determinants of the biogeographical distribution of butterflies in boreal regions

Aim  We explored the relative contributions of climatic and land-cover factors in explaining the distribution patterns of butterflies in a boreal region.
Location  Finland, northern Europe.
Methods  Data from a national butterfly atlas survey carried out during 1991–2003, with a 10-km grain grid system, were used in these analyses. We used generalized additive models (GAM) and hierarchical partitioning (HP) to explore the main environmental correlates (climate and land-cover) of the realized niches of 98 butterfly species. The accuracy of the distribution models (GAMs) was validated by resubstitution and cross-validation approaches, using the area under the curve (AUC) derived from the receiver operating characteristic (ROC) plots.
Results  Predictive accuracies of the 98 individual environment–butterfly models varied from low to very high (cross-validated AUC values 0.48–0.99), with a mean of 0.79. The results of both the GAM and HP analyses were broadly concordant. Most of the variation in butterfly distributions is associated with growing degree-days, mean temperature of the coldest month and cover of built-up area in all six phylogenetic groups (butterfly families). There were no statistically significant differences in predictive accuracy among the different butterfly families.
Main conclusions  About three-quarters of the distributions of butterfly species in Finland appear to be governed principally by climatic, predominantly temperature-related, factors. This indicates that many butterfly species may respond rapidly to the projected climate change in boreal regions. By determining the ecological niches of multiple species, we can project their range shifts in response to changes in climate and land-cover, and identify species that are particularly sensitive to forecasted global changes.     

高尔基体堆形成的分子机制

高尔基体堆(golgi stack)的形成对高尔基体行使功能起着至关重要的作用。体外的无细胞实验系统已经鉴定了很多在高尔基堆形成中起作用的蛋白,并总结了它们起作用的模式。NSF和p97能分别介导有丝分裂后的扁平囊(cisterna)重生。依赖于NSF的扁平囊重生必需“栓链(tether)”giantin- p115-GM130的作用,该“栓链”也在随后的扁平囊堆叠中起作用。扁平囊的堆叠主要依赖GRASP65和GRASP55的相互作用。哺乳动物细胞中,高尔基堆层通过侧向连接形成高尔基带(golgi ribbon)。GM130和GRASP65对高尔基带(golgi ribbon)形成是必需的。     

A spatio‐temporal framework for efficient inventories of natural resources: A case study with submersed macrophytes

Questions: Does a reduced nutrient load in open water increase species richness and the importance of regional and local site characteristics for species abundance and spatial distribution? Can we build lake‐specific models of macrophyte abundance and distribution based on site characteristics in order to prepare a cost‐efficient framework for future surveys? Location: Lake Constance, 47°39′N, 9°18′E. Methods: Generalized additive models (GAMs) were used to predict the potential distributions of eight species and overall species richness. Submersed macrophyte distribution in 1993 was compared with corresponding data from 1978, when eutrophication was at its maximum. Results: Spatial predictions for eight species and overall species richness were relatively accurate and independent of water chemistry. Depth was confirmed as a main predictor of species distribution, while effective fetch distance was retained in many models. Mineralogical variables of sediment composition represent allogenic and autogenic sediment sources and their east‐west gradient in Lake Constance corresponded to east‐west gradients of species distribution and richness. GAMs appeared more efficient than generalized linear models (GLMs) for modelling species responses to environmental gradients. Conclusions: Reduced trophic status increases species richness and the importance of regional and local site characteristics for species abundance and distribution. Our models represent a spatio‐temporal framework for future lake monitoring purposes and allow the development of effective monitoring; this could be generalized for many ecosystem types and would be particularly efficient for large lakes such as Lake Constance.     

Monomerization and ER Relocalization of GRASP Is a Requisite for Unconventional Secretion of CFTR

Induction of endoplasmic reticulum (ER)‐to‐Golgi blockade or ER stress induces Golgi reassembly stacking protein (GRASP)‐mediated, Golgi‐independent unconventional cell‐surface trafficking of the folding‐deficient ΔF508‐cystic fibrosis transmembrane conductance regulator (CFTR). However, molecular mechanisms underlying this process remain elusive. Here, we show that phosphorylation‐dependent dissociation of GRASP homotypic complexes and subsequent relocalization of GRASP to the ER play a critical role in the unconventional secretion of CFTR. Immunolocalization analyses of mammalian cells revealed that the Golgi protein GRASP55 was redistributed to the ER by stimuli that induce unconventional secretion of ΔF508‐CFTR, such as induction of ER‐to‐Golgi blockade by the Arf1 mutant. Notably, the same stimuli also induced phosphorylation of regions near the C‐terminus of GRASP55 and dissociation of GRASP homomultimer complexes. Furthermore, phosphorylation‐mimicking mutations of GRASP55 induced the monomerization and ER relocalization of GRASP55, and these changes were nullified by phosphorylation‐inhibiting mutations. These results provide mechanistic insights into how GRASP accesses the ER‐retained ΔF508‐CFTR and mediates the ER stress‐induced unconventional secretion pathway.      

Sorting nexin 27 interacts with the Cytohesin associated scaffolding protein (CASP) in lymphocytes

CASP is a small cytokine-inducible protein, primarily expressed in hematopoetic cells, which associates with members of the Cytohesin/ARNO family of guanine nucleotide-exchange factors. Cytohesins activate ARFs, a group of GTPases involved in vesicular initiation. Functionally, CASP is an adaptor protein containing a PDZ domain, a coiled-coil, and a potential carboxy terminal PDZ-binding motif that we sought to characterize here. Using GST pulldowns and mass spectrometry we identified the novel interaction of CASP and sorting nexin 27 (SNX27). In lymphocytes, CASP's PDZ-binding motif interacts with the PDZ domain of SNX27. This protein is a unique member of the sorting nexin family of proteins, a group generally involved in the endocytic and intracellular sorting machinery. Endogenous SNX27 and CASP co-localize at the early endosomal compartment in lymphocytes and also in transfection studies. These results suggest that endosomal SNX27 may recruit CASP to orchestrate intracellular trafficking and/or signaling complexes.     

Evolution and diversity of the Golgi body

Often considered a defining eukaryotic feature, the Golgi body is one of the most recognizable and functionally integrated cellular organelles. It is therefore surprising that some unicellular eukaryotes do not, at first glance, appear to possess Golgi stacks. Here we review the molecular evolutionary, genomic and cell biological evidence for Golgi bodies in these organisms, with the organelle likely present in some form in all cases. This, along with the overwhelming prevalence of stacked cisternae in most eukaryotes, implies that the ancestral eukaryote possessed a stacked Golgi body, with at least eight independent instances of Golgi unstacking in our cellular history.