Voltage-gated sodium channels potentiate the invasive capacities of human non-small-cell lung cancer cell lines.

Ionic channel activity is involved in fundamental cellular behaviour and participates in cancerous features such as proliferation, migration and invasion which in turn contribute to the metastatic process. In this study, we investigated the expression and role of voltage-gated sodium channels in non-small-cell lung cancer cell lines. Functional voltage-gated sodium channels expression was investigated in normal and non-small-cell lung cancer cell lines. The measurement, in patch-clamp conditions, of tetrodotoxin-inhibitable sodium currents indicated that the strongly metastatic cancerous cell lines H23, H460 and Calu-1 possess functional sodium channels while normal and weakly metastatic cell lines do not. While all the cell lines expressed mRNA for numerous sodium channel isoforms, only H23, H460 and Calu-1 cells had a 250 kDa protein corresponding to the functional channel. The other cell lines also had another protein of 230 kDa which is not addressed to the membrane and might act as a dominant negative isoform to prevent channel activation. At the membrane potential of these cells, channels are partially open. This leads to a continuous entry of sodium, disrupting sodium homeostasis and down-stream signaling pathways. Inhibition of the channels by tetrodotoxin was responsible for a 40-50% reduction of in vitro invasion. These experiments suggest that the functional expression of voltage-gated sodium channels might be an integral component of the metastatic process in non-small-cell lung cancer cells probably through its involvement in the regulation of intracellular sodium homeostasis. These channels could serve both as novel markers of the metastatic phenotype and as potential new therapeutic targets

HOG1 AND THE HOMEOSTASIS OF TELOMERES DURING REPLICATIVE SENESCENCE IN SACCHAROMYCES CEREVISIAE

Telomeres protect linear chromosomes from fusions and degradations. However, the protective functions of telomeres are put at risk when they shorten as the cell divides without telomerase. When telomeres reach a critically short length, they trigger a DNA damage checkpoint-dependent cell cycle arrest in a process called replicative senescence. Data obtained in different organisms show that mitochondrial defects and oxidative stress accelerate telomere shortening and dysfunction. However, how ROS levels are produced and controlled during senescence is still elusive. In Saccharomyces cerevisiae, Hog1, the orthologue of p38 MAPK, is a major factor protecting against osmotic stress and regulating ROS levels. Hog1 activates the transcription of detoxifying genes but also interferes with mitochondrial functions and increases the levels of ROS. Here we show that in telomerase-negative cultures, ROS levels increase before senescence crisis, coinciding with the activation of Hog1. Deletion of Hog1 accelerates senescence demonstrating its contribution to cell viability in the absence of telomerase. Adding antioxidants delays senescence and partially suppresses the acceleration of senescence in hog1-delta cells, suggesting a role of Hog1 in regulating ROS levels during this process. ROS levels also increase during senescence when Mec1, the major DNA damage checkpoint in budding yeast, is mutated. However, Mec1 and Hog1 act on ROS through two independent signaling pathways that cooperate to keep ROS in check during senescence

Hog1 acts in a Mec1-independent manner to counteract oxidative stress following telomerase inactivation in Saccharomyces cerevisiae

Abstract Replicative senescence is triggered when telomeres reach critically short length and activate permanent DNA damage checkpoint-dependent cell cycle arrest. Mitochondrial dysfunction and increase in oxidative stress are both features of replicative senescence in mammalian cells. However, how reactive oxygen species levels are controlled during senescence is elusive. Here, we show that reactive oxygen species levels increase in the telomerase-negative cells of Saccharomyces cerevisiae during replicative senescence, and that this coincides with the activation of Hog1, a mammalian p38 MAPK ortholog. Hog1 counteracts increased ROS levels during replicative senescence. While Hog1 deletion accelerates replicative senescence, we found this could stem from a reduced cell viability prior to telomerase inactivation. ROS levels also increase upon telomerase inactivation when Mec1, the yeast ortholog of ATR, is mutated, suggesting that oxidative stress is not simply a consequence of DNA damage checkpoint activation in budding yeast. We speculate that oxidative stress is a conserved hallmark of telomerase-negative eukaryote cells, and that its sources and consequences can be dissected in S. cerevisiae

Initiation of DNA double strand break repair: signaling and single-stranded resection dictate the choice between homologous recombination, non-homologous end-joining and alternative end-joining

A DNA double strand break (DSB) is a highly toxic lesion, which can generate genetic instability and profound genome rearrangements. However, DSBs are required to generate diversity during physiological processes such as meiosis or the establishment of the immune repertoire. Thus, the precise regulation of a complex network of processes is necessary for the maintenance of genomic stability, allowing genetic diversity but protecting against genetic instability and its consequences on oncogenesis. Two main strategies are employed for DSB repair: homologous recombination (HR) and non-homologous end-joining (NHEJ). HR is initiated by single-stranded DNA (ssDNA) resection and requires sequence homology with an intact partner, while NHEJ requires neither resection at initiation nor a homologous partner. Thus, resection is an pivotal step at DSB repair initiation, driving the choice of the DSB repair pathway employed. However, an alternative end-joining (A-EJ) pathway, which is highly mutagenic, has recently been described; A-EJ is initiated by ssDNA resection but does not require a homologous partner. The choice of the appropriate DSB repair system, for instance according the cell cycle stage, is essential for genome stability maintenance. In this context, controlling the initial events of DSB repair is thus an essential step that may be irreversible, and the wrong decision should lead to dramatic consequences. Here, we first present the main DSB repair mechanisms and then discuss the importance of the choice of the appropriate DSB repair pathway according to the cell cycle phase. In a third section, we present the early steps of DSB repair i.e., DSB signaling, chromatin remodeling, and the regulation of ssDNA resection. In the last part, we discuss the competition between the different DSB repair mechanisms. Finally, we conclude with the importance of the fine tuning of this network for genome stability maintenance and for tumor protection in fine

Initiation of DNA double strand break repair: signaling and single-stranded resection dictate the choice between homologous recombination, non-homologous end-joining and alternative end-joining.

International audienceA DNA double strand break (DSB) is a highly toxic lesion, which can generate genetic instability and profound genome rearrangements. However, DSBs are required to generate diversity during physiological processes such as meiosis or the establishment of the immune repertoire. Thus, the precise regulation of a complex network of processes is necessary for the maintenance of genomic stability, allowing genetic diversity but protecting against genetic instability and its consequences on oncogenesis. Two main strategies are employed for DSB repair: homologous recombination (HR) and non-homologous end-joining (NHEJ). HR is initiated by single-stranded DNA (ssDNA) resection and requires sequence homology with an intact partner, while NHEJ requires neither resection at initiation nor a homologous partner. Thus, resection is an pivotal step at DSB repair initiation, driving the choice of the DSB repair pathway employed. However, an alternative end-joining (A-EJ) pathway, which is highly mutagenic, has recently been described; A-EJ is initiated by ssDNA resection but does not require a homologous partner. The choice of the appropriate DSB repair system, for instance according the cell cycle stage, is essential for genome stability maintenance. In this context, controlling the initial events of DSB repair is thus an essential step that may be irreversible, and the wrong decision should lead to dramatic consequences. Here, we first present the main DSB repair mechanisms and then discuss the importance of the choice of the appropriate DSB repair pathway according to the cell cycle phase. In a third section, we present the early steps of DSB repair i.e., DSB signaling, chromatin remodeling, and the regulation of ssDNA resection. In the last part, we discuss the competition between the different DSB repair mechanisms. Finally, we conclude with the importance of the fine tuning of this network for genome stability maintenance and for tumor protection in fine

Acide linoléique conjugué et cancer du sein

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Increased BRCA1 protein in mammary tumours of rats fed marine omega-3 fatty acids.

Any factor affecting BRCA gene regulation may be of interest in the prevention of breast tumourigenesis. We studied the influence of dietary docosahexaenoic acid (DHA), a major omega-3 fatty acid present in marine products, on rat autochthonous mammary tumourigenesis. DHA-supplementation significantly reduced the incidence of tumours (30%, P=0.007) and led to a 60% increase (P=0.02) in BRCA1 protein level. Since DHA influences the product of a major tumour suppressor gene, this finding may contribute to the observation that high-fish consumption reduces the risk of breast cancer

A Role for BLM in Double-Strand Break Repair Pathway Choice: Prevention of CtIP/Mre11-Mediated Alternative Nonhomologous End-Joining

The choice of the appropriate double-strand break (DSB) repair pathway is essential for the maintenance of genomic stability. Here, we show that the Bloom syndrome gene product, BLM, counteracts CtIP/MRE11-dependent long-range deletions (>200 bp) generated by alternative end-joining (A-EJ). BLM represses A-EJ in an epistatic manner with 53BP1 and RIF1 and is required for ionizing-radiation-induced 53BP1 focus assembly. Conversely, in the absence of 53BP1 or RIF1, BLM promotes formation of A-EJ long deletions, consistent with a role for BLM in DSB end resection. These data highlight a dual role for BLM that influences the DSB repair pathway choice: (1) protection against CtIP/MRE11 long-range deletions associated with A-EJ and (2) promotion of DNA resection. These antagonist roles can be regulated, according to cell-cycle stage, by interacting partners such as 53BP1 and TopIII, to avoid unscheduled resection that might jeopardize genome integrity

Tissue factor pathway inhibitor 2 is a potent kallikrein-related protease 12 inhibitor

International audienceAbstract The protease activities are tightly regulated by inhibitors and dysregulation contribute to pathological processes such as cancer and inflammatory disorders. Tissue factor pathway inhibitor 2 (TFPI-2) is a serine proteases inhibitor, that mainly inhibits plasmin. This protease activated matrix metalloproteases (MMPs) and degraded extracellular matrix. Other serine proteases are implicated in these mechanisms like kallikreins (KLKs). In this study, we identified for the first time that TFPI-2 is a potent inhibitor of KLK5 and 12. Computer modeling showed that the first Kunitz domain of TFPI-2 could interact with residues of KLK12 near the catalytic triad. Furthermore, like plasmin, KLK12 was able to activate proMMP-1 and -3, with no effect on proMMP-9. Thus, the inhibition of KLK12 by TFPI-2 greatly reduced the cascade activation of these MMPs and the cleavage of cysteine-rich 61, a matrix signaling protein. Moreover, when TFPI-2 bound to extracellular matrix, its classical localisation, the KLK12 inhibition was retained. Finally, TFPI-2 was downregulated in human non-small-cell lung tumour tissue as compared with non-affected lung tissue. These data suggest that TFPI-2 is a potent inhibitor of KLK12 and could regulate matrix remodeling and cancer progression mediated by KLK12