Furin Prodomain ppFurin Enhances Ca2+ Entry Through Orai and TRPC6 Channels’ Activation in Breast Cancer Cells

The intracellular calcium concentration ([Ca2+]i) modulation plays a key role in the regulation of cellular growth and survival in normal cells and failure of [Ca2+]i homeostasis is involved in tumor initiation and progression. Here we showed that inhibition of Furin by its naturally occurring inhibitor the prodomain ppFurin in the MDA-MB-231 breast cancer cells resulted in enhanced store-operated calcium entry (SOCE) and reduced the cell malignant phenotype. Expression of ppFurin in a stable manner in MDA-MB-231 and the melanoma MDA-MB-435 cell lines inhibits Furin activity as assessed by in vitro digestion assays. Accordingly, cell transfection experiments revealed that the ppFurin-expressing cells are unable to adequately process the proprotein convertase (PC) substrates vascular endothelial growth factor C (proVEGF-C) and insulin-like growth factor-1 receptor (proIGF-1R). Compared to MDA-MB-435 cells, expression of ppFurin in MDA-MB-231 and BT20 cells significantly enhanced SOCE and induced constitutive Ca2+ entry. The enhanced SOCE is impaired by inhibition of Orai channels while the constitutive Ca2+ entry is attenuated by silencing or inhibition of TRPC6 or inhibition of Orai channels. Analysis of TRPC6 activation revealed its upregulated tyrosine phosphorylation in ppFurin-expressing MDA-MB-231 cells. In addition, while ppFurin had no effect on MDA-MB-435 cell viability, in MDA-MB-231 cells ppFurin expression reduced their viability and ability to migrate and enhanced their sensitization to the apoptosis inducer hydrogen peroxide and similar results were observed in BT20 cells. These findings suggest that Furin inhibition by ppFurin may be a useful strategy to interfere with Ca2+ mobilization, leading to breast cancer cells’ malignant phenotype repression and reduction of their resistance to treatments.This research was funded by SIRIC-Brio, La Ligue Contre le Cancer and Region Nouvelle Aquitaine to A.M.K. and by PID2019-104084GB-C21 and PID2019-104084GB-C22/AEI/10.13039/501100011033, MICINN (Grant BFU2016-74932-C2) and Junta de Extremadura-Fondo Europeo de Desarrollo Regional (FEDER; Grants IB16046 and GR18061) to J.A.R. and T.S. J.J.L. is supported by a contract from Junta de Extremadura (TA18011). C.C. is supported by a contract from Junta de Extremadura—FEDER

Understanding and simulating the material behavior during multi-particle irradiations

A number of studies have suggested that the irradiation behavior and damage processes occurring during sequential and simultaneous particle irradiations can significantly differ. Currently, there is no definite answer as to why and when such differences are seen. Additionally, the conventional multi-particle irradiation facilities cannot correctly reproduce the complex irradiation scenarios experienced in a number of environments like space and nuclear reactors. Therefore, a better understanding of multi-particle irradiation problems and possible alternatives are needed. This study shows ionization induced thermal spike and defect recovery during sequential and simultaneous ion irradiation of amorphous silica. The simultaneous irradiation scenario is shown to be equivalent to multiple small sequential irradiation scenarios containing latent damage formation and recovery mechanisms. The results highlight the absence of any new damage mechanism and time-space correlation between various damage events during simultaneous irradiation of amorphous silica. This offers a new and convenient way to simulate and understand complex multi-particle irradiation problems

The Ponto-Caspian basin as a final trap for southeastern Scandinavian Ice-Sheet meltwater

This paper provides new data on the evolution of the Caspian Sea and Black Sea from the Last Glacial Maximum until ca. 12 cal kyr BP. We present new analyses (clay mineralogy, grain-size, Nd isotopes and pollen) applied to sediments from the river terraces in the lower Volga, from the middle Caspian Sea and from the western part of the Black Sea. The results show that during the last deglaciation, the Ponto-Caspian basin collected meltwater and fine-grained sediment from the southern margin of the Scandinavian Ice Sheet (SIS) via the Dniepr and Volga Rivers. It induced the deposition of characteristic red-brownish/chocolate-coloured illite-rich sediments (Red Layers in the Black Sea and Chocolate Clays in the Caspian Sea) that originated from the Baltic Shield area according to Nd data. This general evolution, common to both seas was nevertheless differentiated over time due to the specificities of their catchment areas and due to the movement of the southern margin of the SIS. Our results indicate that in the eastern part of the East European Plain, the meltwater from the SIS margin supplied the Caspian Sea during the deglaciation until ∼13.8 cal kyr BP, and possibly from the LGM. That led to the Early Khvalynian transgressive stage(s) and Chocolate Clays deposition in the now-emerged northern flat part of the Caspian Sea (river terraces in the modern lower Volga) and in its middle basin. In the western part of the East European Plain, our results confirm the release of meltwater from the SIS margin into the Black Sea that occurred between 17.2 and 15.7 cal kyr BP, as previously proposed. Indeed, recent findings concerning the evolution of the southern margin of the SIS and the Black Sea, show that during the last deglaciation, occurred a westward release of meltwater into the North Atlantic (between ca. 20 and 16.7 cal kyr BP), and a southward one into the Black Sea (between 17.2 and 15.7 cal kyr BP). After the Red Layers/Chocolate Clays deposition in both seas and until 12 cal kyr BP, smectite became the dominant clay mineral. The East European Plain is clearly identified as the source for smectite in the Caspian Sea sediments. In the Black Sea, smectite originated either from the East European Plain or from the Danube River catchment. Previous studies consider smectite as being only of Anatolian origin. However, our results highlight both, the European source for smectite and the impact of this source on the depositional environment of the Black Sea during considered period

Many Ribosomal Protein Genes Are Cancer Genes in Zebrafish

We have generated several hundred lines of zebrafish (Danio rerio), each heterozygous for a recessive embryonic lethal mutation. Since many tumor suppressor genes are recessive lethals, we screened our colony for lines that display early mortality and/or gross evidence of tumors. We identified 12 lines with elevated cancer incidence. Fish from these lines develop malignant peripheral nerve sheath tumors, and in some cases also other tumor types, with moderate to very high frequencies. Surprisingly, 11 of the 12 lines were each heterozygous for a mutation in a different ribosomal protein (RP) gene, while one line was heterozygous for a mutation in a zebrafish paralog of the human and mouse tumor suppressor gene, neurofibromatosis type 2. Our findings suggest that many RP genes may act as haploinsufficient tumor suppressors in fish. Many RP genes might also be cancer genes in humans, where their role in tumorigenesis could easily have escaped detection up to now

Non-parametric class completeness estimators for collaborative knowledge graphs — the case of wikidata

Collaborative Knowledge Graph platforms allow humans and automated scripts to collaborate in creating, updating and interlinking entities and facts. To ensure both the completeness of the data as well as a uniform coverage of the different topics, it is crucial to identify underrepresented classes in the Knowledge Graph. In this paper, we tackle this problem by developing statistical techniques for class cardinality estimation in collaborative Knowledge Graph platforms. Our method is able to estimate the completeness of a class—as defined by a schema or ontology—hence can be used to answer questions such as “Does the knowledge base have a complete list of all {Beer Brands—Volcanos—Video Game Consoles}?” As a use-case, we focus on Wikidata, which poses unique challenges in terms of the size of its ontology, the number of users actively populating its graph, and its extremely dynamic nature. Our techniques are derived from species estimation and data-management methodologies, and are applied to the case of graphs and collaborative editing. In our empirical evaluation, we observe that i) the number and frequency of unique class instances drastically influence the performance of an estimator, ii) bursts of inserts cause some estimators to overestimate the true size of the class if they are not properly handled, and iii) one can effectively measure the convergence of a class towards its true size by considering the stability of an estimator against the number of available instances

The Spider Effect: Morphological and Orienting Classification of Microglia in Response to Stimuli in Vivo

The different morphological stages of microglial activation have not yet been described in detail. We transected the olfactory bulb of rats and examined the activation of the microglial system histologically. Six stages of bidirectional microglial activation (A) and deactivation (R) were observed: from stage 1A to 6A, the cell body size increased, the cell process number decreased, and the cell processes retracted and thickened, orienting toward the direction of the injury site; until stage 6A, when all processes disappeared. In contrast, in deactivation stages 6R to 1R, the microglia returned to the original site exhibiting a stepwise retransformation to the original morphology. Thin highly branched processes re-formed in stage 1R, similar to those in stage 1A. This reverse transformation mirrored the forward transformation except in stages 6R to 1R: cells showed multiple nuclei which were slowly absorbed. Our findings support a morphologically defined stepwise activation and deactivation of microglia cells

Changes in calcium dynamics following the reversal of the sodium-calcium exchanger have a key role in AMPA receptor-mediated neurodegeneration via calpain activation in hippocampal neurons

Proteolytic cleavage of the Na(+)/Ca(2+) exchanger (NCX) by calpains impairs calcium homeostasis, leading to a delayed calcium overload and excitotoxic cell death. However, it is not known whether reversal of the exchanger contributes to activate calpains and trigger neuronal death. We investigated the role of the reversal of the NCX in Ca(2+) dynamics, calpain activation and cell viability, in alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptor-stimulated hippocampal neurons. Selective overactivation of AMPA receptors caused the reversal of the NCX, which accounted for approximately 30% of the rise in intracellular free calcium concentration ([Ca(2+)](i)). The NCX reverse-mode inhibitor, 2-[2-[4-(4-nitrobenzyloxy)phenyl]ethyl]isothiourea (KB-R7943), partially inhibited the initial increase in [Ca(2+)](i), and prevented a delayed increase in [Ca(2+)](i). In parallel, overactivation of AMPA receptors strongly activated calpains and led to the proteolysis of NCX3. KB-R7943 prevented calpain activation, cleavage of NCX3 and was neuroprotective. Silencing of NCX3 reduced Ca(2+) uptake, calpain activation and was neuroprotective. Our data show for the first time that NCX reversal is an early event following AMPA receptor stimulation and is linked to the activation of calpains. Since calpain activation subsequently inactivates NCX, causing a secondary Ca(2+) entry, NCX may be viewed as a new suicide substrate operating in a Ca(2+)-dependent loop that triggers cell death and as a target for neuroprotectio

Loss of Ribosomal Protein L11 Affects Zebrafish Embryonic Development through a p53-Dependent Apoptotic Response

Ribosome is responsible for protein synthesis in all organisms and ribosomal proteins (RPs) play important roles in the formation of a functional ribosome. L11 was recently shown to regulate p53 activity through a direct binding with MDM2 and abrogating the MDM2-induced p53 degradation in response to ribosomal stress. However, the studies were performed in cell lines and the significance of this tumor suppressor function of L11 has yet to be explored in animal models. To investigate the effects of the deletion of L11 and its physiological relevance to p53 activity, we knocked down the rpl11 gene in zebrafish and analyzed the p53 response. Contrary to the cell line-based results, our data indicate that an L11 deficiency in a model organism activates the p53 pathway. The L11-deficient embryos (morphants) displayed developmental abnormalities primarily in the brain, leading to embryonic lethality within 6–7 days post fertilization. Extensive apoptosis was observed in the head region of the morphants, thus correlating the morphological defects with apparent cell death. A decrease in total abundance of genes involved in neural patterning of the brain was observed in the morphants, suggesting a reduction in neural progenitor cells. Upregulation of the genes involved in the p53 pathway were observed in the morphants. Simultaneous knockdown of the p53 gene rescued the developmental defects and apoptosis in the morphants. These results suggest that ribosomal dysfunction due to the loss of L11 activates a p53-dependent checkpoint response to prevent improper embryonic development

Genes Involved in the Balance between Neuronal Survival and Death during Inflammation

Glucocorticoids are potent regulators of the innate immune response, and alteration in this inhibitory feedback has detrimental consequences for the neural tissue. This study profiled and investigated functionally candidate genes mediating this switch between cell survival and death during an acute inflammatory reaction subsequent to the absence of glucocorticoid signaling. Oligonucleotide microarray analysis revealed that following lipopolysaccharide (LPS) intracerebral administration at striatum level, more modulated genes presented transcription impairment than exacerbation upon glucocorticoid receptor blockage. Among impaired genes we identified ceruloplasmin (Cp), which plays a key role in iron metabolism and is implicated in a neurodegenative disease. Microglial and endothelial induction of Cp is a natural neuroprotective mechanism during inflammation, because Cp-deficient mice exhibited increased iron accumulation and demyelination when exposed to LPS and neurovascular reactivity to pneumococcal meningitis. This study has identified genes that can play a critical role in programming the innate immune response, helping to clarify the mechanisms leading to protection or damage during inflammatory conditions in the CNS