Caspases Disrupt the Nuclear-Cytoplasmic Barrier

During apoptosis, caspases, a family of proteases, disassemble a cell by cleaving a set of proteins. Caspase-3 plays a major role in the disassembly of the nucleus by processing several nuclear substrates. The question is how caspase-3, which is usually cytoplasmic, gains access to its nuclear targets. It was suggested that caspase-3 is actively transported to the nucleus through the nuclear pores. We found that caspase-9, which is activated earlier than caspase-3, directly or indirectly inactivates nuclear transport and increases the diffusion limit of the nuclear pores. This increase allows caspase-3 and other molecules that could not pass through the nuclear pores in living cells to enter or leave the nucleus during apoptosis by diffusion. Hence, caspase-9 contributes to cell disassembly by disrupting the nuclear-cytoplasmic barrier

Are scientists a workforce? – Or, how Dr. Frankenstein made biomedical research sick

A proposed plan to rescue US biomedical research from its current ‘malaise’ will not be effective as it misdiagnoses the root cause of the diseas

Who is Dr. Frankenstein? Or, what Professor Hayek and his friends have done to science

This commentary suggests that the ongoing malaise of biomedical research results from adopting a doctrine that is incompatible with the principles of creative scientific discovery and thus should be treated as a mental rather than somatic disorder. I overview the progression of the malaise, outline the doctrine and the history of its marriage to science, formulate the diagnosis, justify it by reviewing the symptoms of the malaise, and suggest how to begin to cure the disease

The two cytochrome c species, DC3 and DC4, are not required for caspase activation and apoptosis in Drosophila cells

In Drosophila, activation of the apical caspase DRONC requires the apoptotic protease-activating factor homologue, DARK. However, unlike caspase activation in mammals, DRONC activation is not accompanied by the release of cytochrome c from mitochondria. Drosophila encodes two cytochrome c proteins, Cytc-p (DC4) the predominantly expressed species, and Cytc-d (DC3), which is implicated in caspase activation during spermatogenesis. Here, we report that silencing expression of either or both DC3 and DC4 had no effect on apoptosis or activation of DRONC and DRICE in Drosophila cells. We find that loss of function mutations in dc3 and dc4, do not affect caspase activation during Drosophila development and that ectopic expression of DC3 or DC4 in Drosophila cells does not induce caspase activation. In cell-free studies, recombinant DC3 or DC4 failed to activate caspases in Drosophila cell lysates, but remarkably induced caspase activation in extracts from human cells. Overall, our results argue that DARK-mediated DRONC activation occurs independently of cytochrome c

Deficiency in glutamine but not glucose induces MYC-dependent apoptosis in human cells

The idea that conversion of glucose to ATP is an attractive target for cancer therapy has been supported in part by the observation that glucose deprivation induces apoptosis in rodent cells transduced with the proto-oncogene MYC, but not in the parental line. Here, we found that depletion of glucose killed normal human cells irrespective of induced MYC activity and by a mechanism different from apoptosis. However, depletion of glutamine, another major nutrient consumed by cancer cells, induced apoptosis depending on MYC activity. This apoptosis was preceded by depletion of the Krebs cycle intermediates, was prevented by two Krebs cycle substrates, but was unrelated to ATP synthesis or several other reported consequences of glutamine starvation. Our results suggest that the fate of normal human cells should be considered in evaluating nutrient deprivation as a strategy for cancer therapy, and that understanding how glutamine metabolism is linked to cell viability might provide new approaches for treatment of cancer

Mitotic phosphatases: no longer silent partners

Recent work has highlighted the important role played by protein phosphatase complexes in the regulation of mitosis from yeast to mammals. There have been important advances in defining the roles of the protein serine/threonine phosphatases PP1 and PP2A and the dual specificity protein tyrosine phosphatases CDC25 and Cdc14. Three independent studies defined a regulatory role for PP2A in the control of sister chromatid cohesion, involving a direct interaction with shugoshin. A chromatin targeting subunit has been identified for PP1 and the complex shown to play an essential role in chromosome segregation. Key regulatory residues within CDC25 have been mapped and its activity tied both to the initial activation of cyclin-dependent kinases at the centrosome and to DNA damage checkpoints. Novel roles have been defined for Cdc14, including regulation of rDNA and telomere segregation and participation in spindle assembly. These exciting advances show that protein phosphatases are not merely silent partners to kinases in regulating the control of cell division. Introduction The process of cell division is complex and involves multiple independent regulatory steps, most of which are controlled by reversible protein phosphorylation. In higher eukaryotes, mitosis involves condensation of chromosomes, disassembly of the nuclear lamina, breakdown of the nuclear envelope and disassembly of many forms of nuclear bodies, including nucleoli. Completion of mitosis requires alignment and proper segregation of chromosomes into daughter cells followed by reassembly of nuclei and cytokinesis. These and many other events, such as centrosome separation and spindle assembly, are tightly regulated, and several critical checkpoints occur during mitosis to ensure fidelity. Failure to complete any of the key steps can trigger apoptosis and cell death. While the important role of protein phosphorylation in regulating mitotic events has long been recognized, much of the work in this area has focused on the kinases, primarily the Cdk/Cyclin, Aurora, Polo-like and NIMA families (see This review will focus on recent advances in understanding the contributions of four major classes of protein phosphatases to the regulation of processes involved in controlling cell division, specifically the protein serine/ threonine phosphatases PP1 and PP2A and the dualspecificity protein tyrosine phosphatases (DUSPs) CDC25 and Cdc14. We will draw on examples from species as diverse as yeast, insects and mammals, reflecting the high evolutionary conservation of these regulated events. Serine/threonine phosphatases Both PP1 (termed Glc7 in budding yeast and Dis2 in fission yeast) and PP2A are serine/threonine-specific protein phosphatase catalytic subunits that form holoenzyme complexes with one or more regulatory subunits. These regulatory subunits can affect cellular location and/or substrate specificity. In contrast with most kinases, the PP1 and PP2A catalytic subunits can potentially act on a wide range of substrates and thus substrate specificity is conferred by their interaction partners. Thus, a critical step in understanding the role of PP1 and PP2A holoenzymes is to define their regulatory subunits and the mechanism by which they are targeted to their physiological substrates. Much of the literature ascribing specific roles to PP1 or PP2A has relied on differential effects of inhibitors such as okadaic acid, which in vitro blocks PP2A activity at lower concentrations than are required to inhibit PP1 PP2A PP2A plays a prominent role in the regulation of mitosis and signalling pathways. In addition to its interaction with both scaffolding and variable subunits (termed 'A' and 'B' subunits, respectively) in a trimeric complex (see Using immunoprecipitation and yeast two-hybrid studies, several groups independently identified a specific PP2A trimeric complex that interacts with Sgo1 [7 ,8 ,9 ]. On the basis of RNAi studies and analysis of a non-PP2A-binding hSgo1 mutant, Tang and colleagues [7 ] proposed that interaction with PP2A is required for centromeric localization of hSgo1 and proper chromosome segregation. Independently, the same PP2A complex was immunopurified from HEK 293T cells using Flagtagged hSgo1 [8 ]. Immunofluorescence studies by Kitajima and colleagues showed colocalization of hSgo1 and the B56 PP2A regulatory subunit at mammalian centromeres. Using RNAi in mammals, they also reported that knockdown of hSgo2, but not of hSgo1, resulted in loss of centromeric PP2A. Conversely, knockdown of PP2A led to a loss of centromeric hSgo1 [8 ]. Studies on both budding and fission yeast undergoing meiosis also showed that Sgo1 interacts with PP2A at centromeres and serves to protect the cohesin Rec8 subunit from phosphorylation and cleavage [9 ]. Interestingly, tethering of yeast PP2A at specific sites on chromosome arms preserved cohesion at these sites even after meiosis I, when arm cohesin should dissociate, showing an intrinsic ability of PP2A to protect cohesin, independent of Sgo1 [8 ,9 ]. The PP2A complex may thus work both directly at centromeres to maintain cohesion and by facilitating accumulation of Sgo1, which also acts to prevent cleavage of cohesin. Taken together, these studies point to an important new role for PP2A in the control of chromosome cohesion, mediated, at least in part, through interactions with shugoshins ( PP2A has also been implicated in regulating mitotic exit. Wang and Ng [10] provided evidence suggesting that a PP2A-Cdc55 complex dephosphorylates the mitotic exit network (MEN) activator Tem1 in budding yeast. This prevents mitotic exit by blocking release of Cdc14 from 624 Cell division, growth and death Figure 1 Role of PP2A in maintenance of chromosome cohesion. This diagram summarizes three recent studies that identified a specific PP2A trimeric complex acting with shugoshin to protect cohesin at centromeres from phosphorylation and cleavage until the metaphase-anaphase transition. In metazoan mitosis, cohesin is removed from chromosome arms at prometaphase but remains at the centromere regions, protected by shugoshin and PP2A. At the metaphase-anaphase transition, separase is activated and cleaves this residual cohesin, resulting in a loss of cohesion and separation of sister chromatids. PP1 PP1 has been shown to contribute to the regulation of multiple cellular processes including glycogen metabolism and muscle contraction, mediated by interaction of the PP1 catalytic domain with regulatory proteins termed 'targeting subunits'. Over 50 have been described to date, and they have the potential to regulate both the localization and the catalytic activity of PP1 (see [14] for review). Most targeting subunits share a common 'RVXF' motif that mediates direct binding to PP1 An elegant series of experiments has described a role for PP1 in controlling nuclear envelope assembly at the end of mitosis [24][25][26]. When cells enter mitosis, nuclear lamina disassembly is promoted by phosphorylation of B-type lamins. AKAP149, an ER and nuclear membrane protein, was shown to target PP1 (via an RVXF motif) to dephosphorylate B-type lamins at telophase, enabling their polymerization and thus lamina reassembly. A short peptide from AKAP149 containing the RVXF motif can displace PP1 and induce mislocalization of B-type lamins to the cytoplasm. Although the cells were able to complete mitosis, they died by apoptosis within six hours, suggesting that disruption of lamin assembly may directly trigger apoptosis. The association of PP1 isoforms with centrosomes, kinetochores and the cellular cortex and midbody region (see Pinsky et al. [29 ] took advantage of the regulation of Glc7 by targeting subunits to explore its interaction with Ipl1 (Aurora B) in budding yeast. Glc7 is known to antagonize Ipl1 activity, but it was unclear whether it dephosphorylates its substrates or regulates the kinase directly. Although the targeting subunit has not been identified, titratation of Glc7 away from Ipl1 by overexpression of Glc7 binding proteins that do not play roles in chromosome segregation led to increased phosphorylation of Ipl1 substrates, leading the authors to propose that Glc7 acts to ensure accurate chromosome segregation by dephosphorylating Ipl1 targets. CDC25 CDC25 was first identified in fission yeast 20 years ago as a factor required for entry into mitosis [30]. It is now known to activate cyclin-dependent kinases (Cdks) by removing inhibitory phosphates, which leads to Cdk phosphorylation of multiple substrates that drive the cell division process forward (see Three mammalian genes were identified that complement the yeast cdc25 knockout strain. The proteins encoded by these genes, termed CDC25A, CDC25B and CDC25C, are 60% identical in their C-terminal regions, which include the catalytic core containing the CX 5 R motif common to all protein tyrosine phosphatases. In contrast to the reasonably high homology of their catalytic domains, they are only 20-25% identical in their N-terminal regulatory domains, which contain sites for various post-translational modifications and sitespecific protein interactions, including phosphorylation of key serine and threonine residues, ubiquitination, phosphorylation-dependent binding of 14-3-3 proteins and Pin1-dependent prolyl isomerization (see There is a dramatic hyperphosphorylation of CDC25 during the transition from interphase to mitosis, and several mitotic phosphorylation sites have been mapped (see While all three mammalian CDC25 phosphatases activate their Cdk substrates in the same manner, they appear to have distinct roles in regulating cell cycle transitions (see Mitotic phosphatases: no longer silent partners Trinkle-Mulcahy and Lamond 627 have not yet been ruled out. (b) The G 2 /M transition is regulated in a similar way, with CDC25 activating Cdk1/Cyclin B by dephosphorylating critical residues. All three mammalian CDC25 isoforms have been implicated in regulation of this pathway. (c) The initial activation of Cdk1/Cyclin B has been shown to occur at centrosomes as they begin to separate during prophase, and involves the phosphorylation and activation of CDC25B by the Ajuba-Aurora A complex. The divergent N-terminal regulatory domains of the three mammalian CDC25 isoforms contain a variety of regulatory sites, including phosphorylation sites, 14-3-3 binding sites, domains that regulate degradation and nuclear import and export signals. Several of these known and recently described regulatory sites have been summarized here for (d) CDC25A, (e) CDC25B and (f) CDC25C. Cdc14 While Cdc25 is a key regulator of initiation of mitosis (and hence DNA damage checkpoint control), Cdc14 is a key regulator of late mitotic events, coordinating the temporal and spatial control of chromosome segregation with mitotic spindle disassembly and cytokinesis. In the budding yeast S. cerevisiae, Cdc14p plays a key role in exit from mitosis by dephosphorylating Cdk targets (reviewed in FEAR-controlled release of Cdc14p in budding yeast is also important for division of nucleoli and resolution of highly repetitive rDNA and telomere regions, as demonstrated in two recent studies. These regions separate at mid-anaphase, long after cohesin is cleaved. D'Amours and colleagues 628 Cell division, growth and death Figure 4 Cross-species comparison of Cdc14 localization and function. Cdc14 homologues from four different eukaryotes are listed, showing their localization during interphase and throughout mitosis. Nuclei are shown in green, spindle pole bodies (centrosomes) in red, microtubules in pink and chromosomes in blue. The localization of Cdc14 at these sites is shown in yellow. Known mitotic functions for these homologues are also listed

International Consensus on Guiding Recommendations for Management of Patients with Nonsteroidal Antiinflammatory Drugs Induced Gastropathy-ICON-G

Introduction: Nonsteroidal anti-inflammatory drugs (NSAIDs), one of the most commonly used medications worldwide, are frequently associated with gastrointestinal adverse events. Primary care physicians often face the challenge of achieving adequate pain relief with NSAIDs, while keeping their adverse events to a minimum. This is especially true when long-term use of NSAIDs is required such as in patients with osteoarthritis and rheumatoid arthritis. To help primary care physicians deal with such challenges more effectively, a panel of expert gastroenterologists came together with the aim of developing practice recommendations. Methods: A modified ‘Delphi’ process was used to reach consensus and develop practice recommendations. Twelve gastroenterologists from nine countries provided their expert inputs to formulate the recommendations. These recommendations were carefully developed taking into account existing literature, current practices, and expert opinion of the panelists. Results: The expert panel developed a total of fifteen practice recommendations. Following are the key recommendations: NSAIDs should be prescribed only when necessary; before prescribing NSAIDs, associated modifiable and non-modifiable risk factors should be considered; H. pylori infection should be considered and treated before initiating NSAIDs; patients should be properly educated regarding NSAIDs use; patients who need to be on long-term NSAIDs should be prescribed a gastroprotective agent, preferably a proton pump inhibitor and these patients should be closely monitored for any untoward adverse events. Conclusion/clinical significance: These practice recommendations will serve as an important tool for primary care physicians and will guide them in making appropriate therapeutic choices for their patients. Keywords: Gastropathy, Gastroprotective agents, Non-prescription drugs, Nonsteroidal Anti-inflammatory Agents, Proton pump inhibitor. How to cite this article: Hunt R, Lazebnik LB, Marakhouski YC, Manuc M, Ramesh GN, Aye KS, Bordin DS, Bakulina NV, Iskakov BS, Khamraev AA, Stepanov YM, Ally R, Garg A. International Consensus on Guiding Recommendations for Management of Patients with Nonsteroidal Anti-inflammatory Drugs Induced Gastropathy-ICON-G. Euroasian J Hepatogastroenterol, 2018;8(2):148-160. Source of support: Nil Conflict of interest: Richard Hunt has served as a consultant for INSYS, Dr Reddy's, Takeda, and Novartis. He has received an honorarium from Novartis, Danone, Dr Reddy's, and Takeda. He has been on the speaker's bureau for Takeda and Dr Reddy's and on scientific advisory board for INSYS. Dmitry S Bordin has served as a lecturer for Astellas, AstraZeneca, KRKA and Abbott. For the remaining authors, there are no conflicts of interest

The Problem of Colliding Networks and its Relation to Cancer

Complex systems, ranging from living cells to human societies, can be represented as attractor networks, whose basic property is to exist in one of allowed states, or attractors. We noted that merging two systems that are in distinct attractors creates uncertainty, as the hybrid system cannot assume two attractors at once. As a prototype of this problem, we explore cell fusion, whose ability to combine distinct cells into hybrids was proposed to cause cancer. By simulating cell types as attractors, we find that hybrids are prone to assume spurious attractors, which are emergent and sporadic states of networks, and propose that cell fusion can make a cell cancerous by placing it into normally inaccessible spurious states. We define basic features of hybrid networks and suggest that the problem of colliding networks has general significance in processes represented by attractor networks, including biological, social, and political phenomena