PARP-1 Regulates Metastatic Melanoma through Modulation of Vimentin-induced Malignant Transformation

PARP inhibition can induce anti-neoplastic effects when used as monotherapy or in combination with chemo- or radiotherapy in various tumor settings; however, the basis for the anti-metastasic activities resulting from PARP inhibition remains unknown. PARP inhibitors may also act as modulators of tumor angiogenesis. Proteomic analysis of endothelial cells revealed that vimentin, an intermediary filament involved in angiogenesis and a specific hallmark of EndoMT (endothelial to mesenchymal transition) transformation, was down-regulated following loss of PARP-1 function in endothelial cells. VE-cadherin, an endothelial marker of vascular normalization, was up-regulated in HUVEC treated with PARP inhibitors or following PARP-1 silencing; vimentin over-expression was sufficient to drive to an EndoMT phenotype. In melanoma cells, PARP inhibition reduced pro-metastatic markers, including vasculogenic mimicry. We also demonstrated that vimentin expression was sufficient to induce increased mesenchymal/pro-metastasic phenotypic changes in melanoma cells, including ILK/GSK3-β-dependent E-cadherin down-regulation, Snail1 activation and increased cell motility and migration. In a murine model of metastatic melanoma, PARP inhibition counteracted the ability of melanoma cells to metastasize to the lung. These results suggest that inhibition of PARP interferes with key metastasis-promoting processes, leading to suppression of invasion and colonization of distal organs by aggressive metastatic cells.     

EXO5-DNA structure and BLM interactions direct DNA resection critical for ATR-dependent replication restart

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Highly efficient CRISPR-Cas9-mediated gene knockout in primary human B cells for functional genetic studies of Epstein-Barr virus infection

Gene editing is now routine in all prokaryotic and metazoan cells but has not received much attention in immune cells when the CRISPR-Cas9 technology was introduced in the field of mammalian cell biology less than ten years ago. This versatile technology has been successfully adapted for gene modifications in human myeloid cells and T cells, among others, but applications to human primary B cells have been scarce and limited to activated B cells. This limitation has precluded conclusive studies into cell activation, differentiation or cell cycle control in this cell type. We report on highly efficient, simple and rapid genome engineering in primary resting human B cells using nucleofection of Cas9 ribonucleoprotein complexes, followed by EBV infection or culture on CD40 ligand feeder cells to drive in vitro B cell survival. We provide proof-of-principle of gene editing in quiescent human B cells using two model genes: CD46 and CDKN2A. The latter encodes the cell cycle regulator p16^INK4a which is an important target of Epstein-Barr virus (EBV). Infection of B cells carrying a knockout of CDKN2A with wildtype and EBNA3 oncoprotein mutant strains of EBV allowed us to conclude that EBNA3C controls CDKN2A, the only barrier to B cell proliferation in EBV infected cells. Together, this approach enables efficient targeting of specific gene loci in quiescent human B cells supporting basic research as well as immunotherapeutic strategies.     

Intra-colony variation in the growth of Acropora formosa: extension rates and skeletal structure of white (zooxanthellae-free) and brown-tipped branches

The presence or absence of zooxanthellae near the tip of Acropora formosa branches is correlated with apical skeletal structure and extension rates. White (zooxanthellae-free) tips are lightly calcified, possess thin, widely spaced skeletal elements and bear only a few, poorly developed radial corallites. Brown tips are heavily calcified, possess smaller axial corallites and larger, more numerous radial corallites. White tips exhibit a range of normally distributed extension rates. Brown tips do not extend, but radial growth and internal calcification continue. These processes progressively alter the appearance and density of brown tipped branches. In addition, the axial corallite of brown tips becomes progressively smaller and is eventually indistinguishable from adjacent radial corallites. Although brown and white tips can change from one form to the other, with a corresponding change in extension rate, it is hypothesized that in brown tips with degenerated axial corallites, a new axial corallite must develop before extension can resume. Brown tips predominate in the interior of arborescent colonies, where space for continued extension is limited. They may therefore represent a means of coordination of growth within a colony. Field and experimental evidence suggest that brown tips may develop in response to micro-environmental conditions. White, zooxanthellae-free zones are also characteristic of other branched and plate-forming species, which exhibit rapid extension in a localized region of the colony.     

A statistical approach for detecting genomic aberrations in heterogeneous tumor samples from single nucleotide polymorphism genotyping data

We describe a statistical method for the characterization of genomic aberrations in single nucleotide polymorphism microarray data acquired from cancer genomes. Our approach allows us to model the joint effect of polyploidy, normal DNA contamination and intra-tumour heterogeneity within a single unified Bayesian framework. We demonstrate the efficacy of our method on numerous datasets including laboratory generated mixtures of normal-cancer cell lines and real primary tumours.     

Occurrence and taxonomic significance of ruminate endosperm

Ruminate endosperm is characterized by its uneven and enlarged surface. A list of 58 angiosperm families in which this trait is known to occur is presented. The simultaneous presence of different rumination types in angiosperms and even within single families leads to the conclusion that ruminate endosperm has originated several times in parallel. Therefore, the mere occurrence of rumination does not provide evidence for phylogenetic hypotheses. Nevertheless, rumination features can provide valuable characters for taxonomic purposes, if structural and ontogenetic evidence is considered.