Annexin2 coating the surface of enlargeosomes is needed for their regulated exocytosis

Enlargeosomes are small cytoplasmic vesicles that undergo rapid, Ca2+-dependent exo/endocytosis. The role of the cytoskeleton in these processes was unknown. In PC12-27 cells, microtubule disassembly had little effect on enlargeosomes, whereas microfilament disassembly increased markedly both their resting and stimulated exocytosis, and inhibited their endocytosis. Even at rest enlargeosomes are coated at their cytosolic surface by an actin-associated protein, annexin2, bound by a dual, Ca2+-dependent and Ca2+-independent mechanism. In contrast, the other enlargeosome marker, desmoyokin/Ahnak, is transported across the organelle membrane, apparently by an ABC transporter, and binds to its lumenal face. Annexin2-GFP expression revealed that, upon stimulation, the slow and random enlargeosome movement increases markedly and becomes oriented toward the plasma membrane. After annexin2 downregulation enlargeosome exocytosis induced by both [Ca2+]i rise and cytoskeleton disruption is inhibited, and the NGF-induced differentiation is blocked. Binding of annexin2 to the enlargeosome membrane, the most extensive ever reported (>50% annexin2 bound to approximately 3% of total membrane area), seems therefore to participate in the regulation of their exocytosis.     

Inhibition of mycobacterial arylamine N-acetyltransferase contributes to anti-mycobacterial activity of Warburgia salutaris

In this study, we show that extracts and a purified compound of Warburgia salutaris exhibit anti-mycobacterial activity against Mycobacterium tuberculosis H37Rv and Mycobacterium bovis BCG Pasteur. The extracts did not inhibit growth of Escherichia coli and were not toxic to cultured mammalian macrophage cells at the concentrations at which anti-mycobacterial activity was observed. The extract and pure compound inhibited pure recombinant arylamine N-acetyltransferase (NAT), an enzyme involved in mycobacterial cell wall lipid synthesis. Moreover, neither extract nor pure compound inhibited growth of a strain of M. bovis BCG in which nat has been deleted suggesting that NAT may indeed be a target within the mycobacterial cell. The purified compound is a novel drimane sesquiterpenoid lactone, 11alpha-hydroxycinnamosmolide. These studies show that W. salutaris is a useful source of anti-tubercular compounds for further analysis and supports the hypothesis of a link between NAT inhibition and anti-mycobacterial activity.     

Effect of membrane microheterogeneity and domain size on fluorescence resonance energy transfer

Studies of multicomponent membranes suggest lateral inhomogeneity in the form of membrane domains, but the size of small (nanoscale) domains in situ cannot be determined with current techniques. In this article, we present a model that enables extraction of membrane domain size from time-resolved fluorescence resonance energy transfer (FRET) data. We expand upon a classic approach to the infinite phase separation limit and formulate a model that accounts for the presence of disklike domains of finite dimensions within a two-dimensional infinite planar bilayer. The model was tested against off-lattice Monte Carlo calculations of a model membrane in the liquid-disordered (l(d)) and liquid-ordered (l(o)) coexistence regime. Simulated domain size was varied from 5 to 50 nm, and two fluorophores, preferentially partitioning into opposite phases, were randomly mixed to obtain the simulated time-resolved FRET data. The Monte Carlo data show clear differences in the efficiency of energy transfer as a function of domain size. The model fit of the data yielded good agreement for the domain size, especially in cases where the domain diameter is <20 nm. Thus, data analysis using the proposed model enables measurement of nanoscale membrane domains using time-resolved FRET.     

Evolutionary history predicts high‐impact invasions by herbivorous insects

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The C-terminus of Gi family G-proteins as a determinant of 5-HT(1A) receptor coupling

Using a universal signaling assay employing G-protein chimeras comprising the C-terminal five amino acids of Gi1/2, Gi3, Go, and Gz fused to Gq, the calcium mobilizing G-protein, we explored the role of the C-terminus of Gi family G-proteins as a determinant for 5-HT(1A) receptor functional coupling. Co-expression of the 5-HT(1A) receptor with each of the Gq/Gi family chimeras resulted in a concentration-dependent increase in calcium upon addition of 5-HT, although the coupling efficiency differed dramatically. Gq/Gi3 resulted in the most efficient coupling based on both potency and relative maximum response to 5-HT. Gq/Go also produced efficient coupling in terms of relative 5-HT efficacy (76% of the Gq/Gi3 maximum response), although 5-HT exhibited 4-fold lower agonist potency, and Gq/Gz and Gq/Gi1/2 conferred poor functional coupling. Agonist potencies and relative efficacies determined for a number of 5-HT(1A) receptor agonists using Gq/Gi3 coupling were significantly weaker than those described previously for coupling through the native G-protein. These results indicate the C-terminus of Gi3 as an important determinant for coupling to the 5-HT(1A) receptor, while the reduced functional agonist activities suggest additional motifs participate in receptor/G-protein coupling.     

Cross-talk between macrophages,smooth muscle cells,and endothelial cells in response to cigarette smoke: the effects on MMP2 and 9

The aim of the present study was to analyze the expression of sex-determining region Y-related high mobility group box 4 (SOX4) in non-small cell lung cancer (NSCLC) and its correlation with clinicopathologic characteristics, including the survival of NSCLC patients. To observe initially the expression status of SOX4 in lung squamous cell carcinoma and adenocarcinoma at gene expression omnibus. The expression of SOX4 mRNA and protein was examined in NSCLC tissues and normal lung tissues through real-time PCR and immunohistochemistry. Meanwhile, the relationship of SOX4 expression levels with clinical characteristics of 168 NSCLC patients was analyzed by immunohistochemistry. Univariate and multivariate analyses were performed to determine the association between SOX4 expression and prognosis of NSCLC patients. In our results, SOX4 expression was increased in NSCLC tissues compared with paired normal lung tissues in microarray data (GSE3268). SOX4 mRNA and protein expression were markedly higher in NSCLC tissues than in normal lung tissues (P = 0.001 and P = 0.001, respectively). Using immunohistochemistry, high levels of SOX4 protein were positively correlated with status of differentiated degree (high vs. middle, P = 0.004; high vs. low, P < 0.001), clinical stage (I–II vs. III–IV, P < 0.001), T classification (T1–T2 vs. T3–T4, P = 0.004), N classification (N0–N1 vs. N2–N3, P = 0.002), and M classification (M0 vs. M1, P = 0.011) in NSCLC. Moreover, the higher level of SOX4 expression was markedly correlated with poor overall survival in NSCLC patients (P < 0.001). Multivariate analysis suggested that increased SOX4 expression was a poor independent prognostic predictor for NSCLC patients (P = 0.002). In conclusion, SOX4 plays an important role on NSCLC progression and prognosis and may serve as a convictive prognostic biomarker for NSCLC patients.     

Genetic variation in cell death genes and risk of non-Hodgkin lymphoma

Background

Non-Hodgkin lymphomas are a heterogeneous group of solid tumours that constitute the 5^th highest cause of cancer mortality in the United States and Canada. Poor control of cell death in lymphocytes can lead to autoimmune disease or cancer, making genes involved in programmed cell death of lymphocytes logical candidate genes for lymphoma susceptibility.

Materials and Methods

We tested for genetic association with NHL and NHL subtypes, of SNPs in lymphocyte cell death genes using an established population-based study. 17 candidate genes were chosen based on biological function, with 123 SNPs tested. These included tagSNPs from HapMap and novel SNPs discovered by re-sequencing 47 cases in genes for which SNP representation was judged to be low. The main analysis, which estimated odds ratios by fitting data to an additive logistic regression model, used European ancestry samples that passed quality control measures (569 cases and 547 controls). A two-tiered approach for multiple testing correction was used: correction for number of tests within each gene by permutation-based methodology, followed by correction for the number of genes tested using the false discovery rate.

Results

Variant rs928883, near miR-155, showed an association (OR per A-allele: 2.80 [95% CI: 1.63–4.82]; p_F = 0.027) with marginal zone lymphoma that is significant after correction for multiple testing.

Conclusions

This is the first reported association between a germline polymorphism at a miRNA locus and lymphoma.     

The changing incidence of human papillomavirus-associated oropharyngeal cancer using multiple imputation from 2000 to 2010 at a Comprehensive Cancer Centre

Introduction Human papillomavirus (HPV) is a risk and prognostic factor for oropharyngeal cancer (OPC). Determining whether the incidence of HPV-associated OPC is rising informs health policy. Methods HPV status was ascribed using p16 immunohistochemistry in 683/1474 OPC patients identified from the Princess Margaret Hospital's Cancer Registry (from 2000 to 2010). Missing p16 data was estimated using multiple (n = 100) imputation (MI) and validated using an independent OPC cohort (n = 214). Non-OPC head and neck squamous cell carcinoma (HNSCC) (n = 3262) were also used for time-trend comparison. Regression was used to compare HNSCC subsets and time-trends. The c-index was used to measure the predictive ability of MI. Results The incidence of OPC rose from 23.3% of all HNSCC in 2000 to 31.2% in 2010 (p = 0.002). In the subset of OPC tested for p16, there was no change in p16 positivity over time (p = 0.9). However, p16 testing became more frequent over time (p < 0.0001), but was nonetheless biased, favouring never-smokers [OR 1.87 (95% CI 1.29–2.70)] and tumors of the tonsil [OR 2.30 (1.52–3.47)] or base-of-tongue [OR 1.72 (1.10–2.70)]. These same factors were also associated with p16-positivity [ORs 3.22 (1.27–8.16), 7.26 (3.50–15.1), 5.83 (2.70–12.7), respectively]. Following MI and normalization, the proportion of OPC that was p16-associated rose from 39.8% in 2000 to 65.0% in 2010, p = 0.002, fully explaining the rise in OPC in our patient population. Conclusion The rise in HNSCC referrals seen from 2000 to 2010 at our institution was driven primarily by p16-associated OPC. MI was necessary to derive reliable conclusions when cases with missing data are considerable.