103,588 research outputs found
Alterations of membrane curvature during influenza virus budding
No full textInfluenza A virus belongs to the Orthomyxoviridae family. It is an enveloped virus that contains a segmented and negative-sense RNA genome. Influenza A viruses cause annual epidemics and occasional major pandemics, are a major cause of morbidity and mortality worldwide, and have a significant financial impact on society. Assembly and budding of new viral particles are a complex and multi-step process involving several host and viral factors. Influenza viruses use lipid raft domains in the apical plasma membrane of polarized epithelial cells as sites of budding. Two viral glycoproteins, haemagglutinin and neuraminidase, concentrate in lipid rafts, causing alterations in membrane curvature and initiation of the budding process. Matrix protein 1 (M1), which forms the inner structure of the virion, is then recruited to the site followed by incorporation of the viral ribonucleoproteins and matrix protein 2 (M2). M1 can alter membrane curvature and progress budding, whereas lipid raft-associated M2 stabilizes the site of budding, allowing for proper assembly of the virion. In the later stages of budding, M2 is localized to the neck of the budding virion at the lipid phase boundary, where it causes negative membrane curvature, leading to scission and virion release
How to halve ploidy : lessons from budding yeast meiosis
No full textMaintenance of ploidy in sexually reproducing organisms requires a specialized form of cell division called meiosis that generates genetically diverse haploid gametes from diploid germ cells. Meiotic cells halve their ploidy by undergoing two rounds of nuclear division (meiosis I and II) after a single round of DNA replication. Research in Saccharomyces cerevisiae (budding yeast) has shown that four major deviations from the mitotic cell cycle during meiosis are essential for halving ploidy. The deviations are (1) formation of a link between homologous chromosomes by crossover, (2) monopolar attachment of sister kinetochores during meiosis I, (3) protection of centromeric cohesion during meiosis I, and (4) suppression of DNA replication following exit from meiosis I. In this review we present the current understanding of the above four processes in budding yeast and examine the possible conservation of molecular mechanisms from yeast to humans
The role of influenza neuraminidase transmembrane domain on budding and virus morphology
No full textInfluenza A virus neuraminidase (NA), a type II transmembrane glycoprotein plays a role in the cleavage of sialic acids and facilitating the release of mature virions from the surface of infected cells. NA has also previously been shown to play a role in virion formation during influenza A virus budding, although the exact mechanisms by which NA contributes to influenza virion formation and morphology is currently unknown. Previous research has shown that mutations within the transmembrane domain (TMD) of NA can result in alteration in virion morphology, particularly in the production of filament like influenza virions. In this research project we examined if the TMD does indeed play a role in influenza virus budding and morphology. We utilised both full and partial mutations of the TMD of NA from A/WSN/33, a primarily spherical lab adapted influenza strain, with the TMD of a primarily filamentous strain A/California/09. To evaluate the effects of TMD on the morphology of a primarily spherical strain with that of filamentous strain. This study used a transfection based virus like particle (VLP) system to examine the effects of TMD alterations on morphology, utilising various biochemical and microscopy methods. Our findings show that as previously indicated mutations within the TMD do result in alterations to virion morphology, as well as showing that despite previous theories both NA and NA’s TMD may play a more active role in in budding and morphology than previously though
Tumour budding and CD8+ T-cells: "attackers" and "defenders" in rectal cancer with and without neoadjuvant chemoradiotherapy.
No full textAIM
Tumour budding ("attacker") and CD8+ T-cells ("defender") are recognized as important parameters for risk stratification in colon cancers and combined, may have even stronger clinical impact. Here, we determine the value of tumor budding and CD8+ in rectal cancer patients treated with/without neoadjuvant therapy.
METHODS
Using digital scans of all tumor slides/case, we analysed CD8+ T-cell counts in two patient cohorts: 45 neoadjuvantly-treated and 47 primarily surgically-treated (totaling n=540 slides) after double-staining of the surgical resection specimen for pan-cytokeratin and CD8+. Tumour buds in hotspots were manually counted (area 0.785 mm2 ), CD8+ T-cell counts were analysed both in tumor budding hotspots and densest CD8+ regions throughout the tumor, separately.
RESULTS
In neoadjuvantly treated patients, only tumor budding and not CD8+ T-cells was associated with tumor features, including more advanced ypT (p=0.0062), venous invasion (p=0.002), lymphatic invasion (p=0.0003) and perineural invasion (p=0.0017) as well as higher AJCC tumor regression score (p=0.0035), indicating less tumor response. Overall survival was also worse in patients with high-grade budding in univariate analysis only. In contrast, all three variables, namely tumor budding (p=0.0347), CD8+ T-cells in budding hotspots (p=0.0382) and CD8+ T-cells in densest areas (p=0.0117) were associated with worse (budding) and better (CD8) survival time also in multivariate setting CONCLUSION: In rectal cancer, tumor budding has clinical relevance in both primarily surgically treated patients and in those with neoadjuvantly treated patients, where it characterizes highly aggressive residual disease. CD8+ T-cell counts appear not to have prognostic relevance in the neoadjuvant context
The rim15-Endosulfine-PP2ACdc55 signalling module regulates entry into gametogenesis and quiescence via distinct mechanisms in budding yeast
No full textQuiescence and gametogenesis represent two distinct survival strategies in response to nutrient starvation in budding yeast. Precisely how environmental signals are sensed by yeast cells to trigger quiescence and gametogenesis is not fully understood. A conserved signalling module consisting of Greatwall kinase, Endosulfine and Protein Phosphatase PP2ACdc55 proteins regulates entry into mitosis in Xenopus egg extracts and meiotic maturation in flies. We report here that an analogous signalling module consisting of the serine-threonine kinase Rim15, the Endosulfines Igo1 and Igo2 and the Protein Phosphatase PP2ACdc55, regulates entry into both quiescence and gametogenesis in budding yeast. PP2ACdc55 inhibits entry into gametogenesis and quiescence. Rim15 promotes entry into gametogenesis and quiescence by converting Igo1 into an inhibitor of PP2ACdc55 by phosphorylating at a conserved serine residue. Moreover, we show that the Rim15-Endosulfine-PP2ACdc55 pathway regulates entry into quiescence and gametogenesis by distinct mechanisms. In addition, we show that Igo1 and Igo2 are required for pre-meiotic autophagy but the lack of pre-meiotic autophagy is insufficient to explain the sporulation defect of igo1Δ igo2Δ cells. We propose that the Rim15-Endosulfine-PP2ACdc55 signalling module triggers entry into quiescence and gametogenesis by regulating dephosphorylation of distinct substrates
Tumour Budding/T-cell infiltrates in Colorectal Cancer: Proposal of a Novel Combined Score.
No full textAIMS
The TNM classification system is used for prognostication purposes and to guide patient management. However, in colorectal cancer (CRC), additional markers are needed to stratify prognostic subgroups. From large bodies of research, two promising markers have emerged: Tumour budding and T-cell host response (CD3, CD8 and CD45RO infiltrates). However, attempts to combine these two parameters have been sparse. The aim of this study was to perform an assessment of potential protagonists that could be used in a combined score (Budding/T-cell Score, BTS).
METHODS AND RESULTS
This descriptive, retrospective study was performed on a multi-punch tissue microarray containing material from 345 patients with Stage I-IV CRC. Areas from tumour centre, front and microenvironment were stained for Pancytokeratin/CD3, Pancytokeratin/CD8 and Pancytokeratin/CD45RO. Tumour buds were scored manually and T-cell infiltrates digitally using open-source software (QuPath). Tumour buds, T-cell counts and combined BTS were associated with clinico-pathological features and overall survival (OS). A higher combined BTS score (Buds/CD8, tumour centre) performed better than budding or CD8/CD3 alone in predicting nodal metastases (p<0.0001, OR 1.466, 95%CI: 1.115-1.928). Only higher BTS (Buds/CD3) was significantly associated with poorer OS on multivariate analysis (p= 0.012, HR 1.218, 95%CI: 1.044-1.419).
CONCLUSIONS
Although CD8+/CD3+ T-cells are predictive of tumour biology in CRC, we found a combined BTS to be stronger in predicting survival and certain features with high clinical relevance, such as nodal metastases, in comparison to budding or T-cells alone. Further studies combining T-cell infiltrates and tumour budding are necessary to optimize risk assessment of CRC
A functional link between the actin cytoskeleton and lipid rafts during budding of filamentous influenza virions
No full textMorphogenesis of influenza virus is a poorly understood process that produces two types of enveloped virion: approximately 100-nm spheres and similar diameter filaments that reach 20 microm in length. Spherical particles assemble at plasma membrane lipid rafts in a process independent of microfilaments. The budding site of filamentous virions is hitherto uncharacterised but their formation involves the actin cytoskeleton. We confirm microfilament involvement in filamentous budding and show that after disruption of cortical actin by jasplakinolide, HA, NP, and M1 redistributed around beta-actin clusters to form novel annular membrane structures. HA in filamentous virions and jasplakinolide-induced annuli was detergent insoluble at 4 degrees C. Furthermore, in both cases HA partitioned into low buoyant density detergent-insoluble glycolipid domains, indicating that filamentous virions and annuli contain reorganised lipid rafts. We propose that the actin cytoskeleton is required to maintain the correct organisation of lipid rafts for incorporation into budding viral filaments
N-terminally myristoylated feline foamy virus Gag allows Env-independent budding of sub-viral particles
No full textFoamy viruses (FVs) are distinct retroviruses classified as Spumaretrovirinae in contrast to the other retroviruses, the Orthoretrovirinae. As a unique feature of FVs, Gag is not sufficient for sub-viral particle (SVP) release. In primate and feline FVs (PFV and FFV), particle budding completely depends on the cognate FV Env glycoproteins. It was recently shown that an artificially added N-terminal Gag myristoylation signal (myr-signal) overcomes this restriction in PFV inducing an Orthoretrovirus-like budding phenotype. Here we show that engineered, heterologous N-terminal myr-signals also induce budding of the distantly related FFV Gag. The budding efficiency depends on the myr-signal and its location relative to the N-terminus of Gag. When the first nine amino acid residues of FFV Gag were replaced by known myr-signals, the budding efficiency as determined by the detection of extracellular SVPs was low. In contrast, adding myr-signals to the intact N-terminus of FFV Gag resulted in a more efficient SVP release. Importantly, budding of myr-Gag proteins was sensitive towards inhibition of cellular N-myristoyltransferases. As expected, the addition or insertion of myr-signals that allowed Env-independent budding of FFV SVPs also retargeted Gag to plasma membrane-proximal sites and other intracellular membrane compartments. The data confirm that membrane-targeted FV Gag has the capacity of SVP formation
T-cell infiltrates and tumor budding: promising prognostic factors in the tumor microenvironment of colorectal cancer
No full textThe tumor microenvironment is the battleground of pro- and anti-tumor factors. On the tumor’s side, epithelial–mesenchymal transition (EMT) and tumor budding is the histomorphologic signature of aggressive disease. On the host’s side, T-effector cell infiltrates are the hallmark of an active anti-tumoral immune defense. These morphological features can be measured by the histopathologist and represent important novel histomorphological prognostic factors in colorectal cancer (CRC)
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