Building up the inhibitory synapses

2nonenoneCherubini E; Zacchi PCherubini, E; Zacchi,

Asbestos Fibers Enhance the TMEM16A Channel Activity in Xenopus Oocytes

Background: The interaction of asbestos fibers with target cell membranes is still poorly investigated. Here, we detected and characterized an enhancement of chloride conductance in Xenopus oocyte cell membranes induced by exposure to crocidolite (Croc) asbestos fibers. Methods: A two-microelectrode voltage clamp technique was used to test the effect of Croc fiber suspensions on outward chloride currents evoked by step membrane depolarization. Calcium imaging experiments were also performed to investigate the variation of 'resting' oocyte [Ca2+]i following asbestos exposure. Results: The increase in chloride current after asbestos treatment, was sensitive to [Ca2+]e, and to specific blockers of TMEM16A Ca2+-activated chloride channels, MONNA and Ani9. Furthermore, asbestos treatment elevated the 'resting' [Ca2+]i likelihood by increasing the cell membrane permeability to Ca2 in favor of a tonic activation of TMEME16A channels. Western blot analysis confirmed that TMEME16A protein was endogenously present in the oocyte cell membrane and absorbed by Croc. Conclusion: the TMEM16A channels endogenously expressed by Xenopus oocytes are targets for asbestos fibers and represent a powerful tool for asbestos-membrane interaction studies. Interestingly, TMEM16A channels are highly expressed in many types of tumors, including some asbestos-related cancers, suggesting them, for the first time, as a possible early target of crocidolite-mediated tumorigenic effects on target cell membranes

The Ferroxidase Hephaestin in Lung Cancer: Pathological Significance and Prognostic Value

Hephaestin (HEPH) belongs to a group of exocytoplasmic ferroxidases which contribute to cellular iron homeostasis by favouring its export. Down-regulation of HEPH expression, possibly by stimulating cell proliferation due to an increase in iron availability, has shown to correlate with poor survival in breast cancer. The lung is particularly sensitive to iron-induced oxidative stress, given the high oxygen tension present, however, HEPH distribution in lung cancer and its influence on prognosis have not been investigated yet. In this study we explored the prognostic value of HEPH and its expression pattern in the most prevalent histotypes of lung cancers, namely lung adenocarcinoma and lung squamous cell carcinoma. In silico analyses, based on UALCAN, Gene Expression Profiling Interactive Analysis (GEPIA) and Kaplan–Meier plotter bioinformatics, revealed a significant correlation between higher levels of HEPH expression and favorable prognosis, in both cancer histotypes. Moreover, TIMER web platform showed a statistically significant association between HEPH expression and cell elements belonging to the tumor microenvironment identified as endothelial cells and a subpopulation of cancer-associated fibroblasts, further confirmed by double immunohistochemical labeling with cell type specific markers. Taken together, these data shed a light on the complex mechanisms of local iron handling lung cancer can exploit to support tumorigenesis

Prognostic Implications of the Complement Protein C1q in Gliomas

The contribution of the complement system in the pathophysiology of brain cancers has been recently considered in light of its well-known involvement in carcinogenesis. Complement system represents an important component of the inflammatory response, which acts as a functional bridge between the innate and adaptive immune response. C1q, the first recognition subcomponent of the complement classical pathway, has recently been shown to be involved in a range of pathophysiological functions that are not dependent on complement activation. C1q is expressed in the microenvironment of various types of human tumors, including melanoma, prostate, mesothelioma, and ovarian cancers, where it can exert a protective or a harmful effect on cancer progression. Despite local synthesis of C1q in the central nervous system, the involvement of C1q in glioma pathogenesis has been poorly investigated. We, therefore, performed a bioinformatics analysis, using Oncomine dataset and UALCAN database in order to assess whether the expression of the genes encoding for the three chains of C1q (C1qA, C1qB, and C1qC) could serve as a potential prognostic marker for gliomas. The obtained results were then validated using an independent glioma cohort from the Chinese Glioma Genome Atlas datasets. Our bioinformatics analysis, coupled with immunohistochemistry and fluorescence microscopy, appears to suggest a positive correlation between higher levels of C1q expression and unfavorable prognosis in a diverse grade of gliomas

THE CLINICOPATHOLOGICAL AND PROGNOSTIC SIGNIFICANCES OF C1Q EXPRESSION IN GLIOMAS: A BIOINFORMATICS ANALYSIS

Introduction. The complement system represents an important component of the inflammatory response and acts as a functional bridge between the innate and adaptive immune response. The contribution of the complement component C1q in the pathophysiology of brain cancers has been recently considered in light of its well-known involvement in carcinogenesis. Brain malignancies arise from cells of the CNS and are classified according to the tissue of phylogenetic origin. Gliomas represent the most common and aggressive form of brain tumours in adults. They derive from glial cells that help to support the functions of the other main brain cells type, the neurons (1). These are a heterogeneous group of diseases with multiple subtypes (1, 2). Glioblastoma multiforme (GBM) is the most common and fatal form of a primary brain tumour, accounting for approximately 60% of all glioma cases (3), whereas grade-II and -III gliomas are the second most common type of glioma in adults (~30%) (3). C1q molecule, together with other complement components, can be locally produced within the CNS by microglia and astrocytes, rendering it an attractive player in primary brain tumour development (4). The role of C1q in gliomas microenvironment is still poorly characterized and it is still quite puzzling whether it exerts a beneficial or a harmful activity for cancer progression. In the present study we performed a bioinformatics analysis aimed at investigating if C1q can serve as a potential prognostic marker for gliomas. Methods. The expression levels of C1qA, C1qB and C1qC genes in gliomas were analysed using Oncomine analysis. Available genomics data from The Cancer Genome Atlas project was used for Kaplan–Meier survival analysis to generate survival probability plots, using UALCAN analysis. Results. From the analysis performed on several data- sets using Oncomine, we showed a significantly higher mRNA expression levels for C1qA, C1qB and C1qC chains were detected in gliomas (different histotypes and grades) as compared to normal brain tissue (Fig. 1). We observed a positive correlation between the mRNA expression of C1qA, C1qB and C1qC mRNA poly- peptide chains and the unfavorable prognosis only in gliomas grade-II and -III, where the survival probability is indeed reduced (P <0.05) (Fig. 2). No correlation was observed in glioblastoma multiforme (Fig. 2). By immu- nohistochemical approaches we detected a high depo- sition of C1q in the tumor microenvironment of both in grade-II and -III gliomas and in GBMs examined (Fig. 3a glioma, 3b glioblastoma multiforme; 20x Magnification). Moreover, in double immunocytochemical experiments we demonstrated that CD68 positive infiltrating cells are actively synthesizing C1q in the tumor micro-envi- ronment. CD68 expression is characteristic of tumor- associated macrophages, whose enrichment in glioma has been associated with poor prognosis (5). Conclusion. In our study C1q expression was significantly correlated with poor survival probability in gliomas grade-II and -III while this is not the case for GBM. These data altogether underline how complex, multifaceted and still poorly understood is the role C1q can exert on tumor progression, and how the very same molecule can differentially affect the outcome depending on the biological context it comes to act

The amplifier effect: how Pin1 empowers mutant p53

Mutation of p53 occurs in 15 to 20% of all breast cancers, and with higher frequency in estrogen-receptor negative and high-grade tumors. Certain p53 mutations contribute to malignant transformation not only through loss of wild-type p53 but also through a gain of function of specific p53 mutations. How these hotspot mutations turn p53 from a tumor suppressor into an oncogene had until now remained incompletely understood. In an elegant paper published in the July 12 issue of Cancer Cell, Girardini and colleagues show how Pin1-mediated prolylisomerization, a regulatory mechanism intended by evolution to support p53's function as a guardian of the genome, can go haywire and accelerate malignant transformation when p53 carries a dominant-negative mutation

Prognostic implications of the complement protein C1q in gliomas

Associazione Italiana per la Ricerca sul Cancro (AIRC); Program Innovative Tools for Cancer Risk Assessment and Diagnosis

A high-throughput immobilized bead screen for stable proteins and multi-protein complexes

We describe an in vitro colony screen to identify Escherichia coli expressing soluble proteins and stable, assembled multiprotein complexes. Proteins with an N-terminal 6His tag and C-terminal green fluorescent protein (GFP) S11 tag are fluorescently labeled in cells by complementation with a coexpressed GFP 1–10 fragment. After partial colony lysis, the fluorescent soluble proteins or complexes diffuse through a supporting filtration membrane and are captured on Talon® resin metal affinity beads immobilized in agarose. Images of the fluorescent colonies convey total expression and the level of fluorescence bound to the beads indicates how much protein is soluble. Both pieces of information can be used together when selecting clones. After the assay, colonies can be picked and propagated, eliminating the need to make replica plates. We used the method to screen a DNA fragment library of the human protein p85 and preferentially obtained clones expressing the full-length ‘breakpoint cluster region-homology' and NSH2 domains. The assay also distinguished clones expressing stable multi-protein complexes from those that are unstable due to missing subunits. Clones expressing stable, intact heterotrimeric E.coli YheNML complexes were readily identified in libraries dominated by complexes of YheML missing the N subunit

Pin1 and WWP2 regulate GluR2 Q/R site RNA editing by ADAR2 with opposing effects

While the essential role of the adenosine deaminase ADAR2 in RNA editing is well established, how it is regulated remains largely unknown. Here, the prolyl isomerase Pin1 and the E3 ubiquitin ligase WWP2 are shown to play a role in regulating ADAR2 localisation and stability