Regulation of the CoA Biosynthetic Complex Assembly in Mammalian Cells

Coenzyme A (CoA) is an essential cofactor present in all living cells. Under physiological conditions, CoA mainly functions to generate metabolically active CoA thioesters, which are indispensable for cellular metabolism, the regulation of gene expression, and the biosynthesis of neurotransmitters. When cells are exposed to oxidative or metabolic stress, CoA acts as an important cellular antioxidant that protects protein thiols from overoxidation, and this function is mediated by protein CoAlation. CoA and its derivatives are strictly maintained at levels controlled by nutrients, hormones, metabolites, and cellular stresses. Dysregulation of their biosynthesis and homeostasis has deleterious consequences and has been noted in a range of pathological conditions, including cancer, diabetes, Reye’s syndrome, cardiac hypertrophy, and neurodegeneration. The biochemistry of CoA biosynthesis, which involves five enzymatic steps, has been extensively studied. However, the existence of a CoA biosynthetic complex and the mode of its regulation in mammalian cells are unknown. In this study, we report the assembly of all five enzymes that drive CoA biosynthesis, in HEK293/Pank1β and A549 cells, using the in situ proximity ligation assay. Furthermore, we show that the association of CoA biosynthetic enzymes is strongly upregulated in response to serum starvation and oxidative stress, whereas insulin and growth factor signaling downregulate their assembly

Deep Impact Mission to Tempel 1 Favours New Explosive Cosmogony of Comets

The assumption that short-period (SP) comets are fragments of massive icy envelopes of Ganymede-like bodies saturated by products of ice electrolysis that underwent global explosions provides a plausible explanation of all known manifestations of comets, including the jet character of outflows, the presence of ions in the vicinity of the nucleus, the bursts and splitting of cometary nuclei, etc., with solar radiation initiating burning of the products of electrolysis in the nucleus. As shown persuasively by numerical simulation carried out in hydrodynamic approximation, the shock wave initiated by the Deep Impact (DI) impactor in the cometary ice saturated originally by the electrolysis products 2H2 + O2 is capable of activating under certain conditions exothermal reactions (of the type O2 + H2 + organics = H2O + CO + HCN + other products of incomplete burning of organics including its light and heavy pyrolyzed compounds, soot, etc.), which will slow down shock wave damping (forced detonation) and increase many times the energy release. As a result, the measured energetics of ejections and outflows from the crater have to exceed the DI energetics. Analysis of different clusters of the DI experiment data confirms these conclusions and expectations and thus it favours the planetary origin of comets.Comment: 21 pages incluging 3 figure

Pseudomonas aeruginosa isolates co-incubated with Acanthamoeba castellanii exhibit phenotypes similar to chronic cystic fibrosis isolates

The opportunistic pathogen, Pseudomonas aeruginosa , is ubiquitous in the environment, and in humans is capable of causing acute and chronic infections. P. aeruginosa , when co-incubated with the bacterivorous amoeba, Acanthamoeba castellanii , for extended periods, produced genetic and phenotypic variants. Sequencing of late-stage amoeba-adapted P. aeruginosa isolates demonstrated single nucleotide polymorphisms within genes that encode known virulence factors, and this correlated with a reduction in expression of virulence traits. Virulence towards the nematode, Caenorhabditis elegans , was attenuated in late-stage amoeba-adapted P. aeruginosa compared to early stage amoeba-adapted and non-adapted counterparts. Late-stage amoeba-adapted P. aeruginosa lost competitive fitness compared to non-adapted counterparts when grown in nutrient rich media. However, non-adapted P. aeruginosa were rapidly cleared by amoeba predation, whereas late-stage amoeba-adapted isolates remained in higher numbers 24 h after ingestion by amoeba. In addition, there was reduced uptake by macrophage of amoeba-adapted isolates and reduced uptake by human neutrophils as well as increased survival in the presence of neutrophils. Our findings indicate that the selection imposed by amoeba on P. aeruginosa resulted in reduced virulence over time. Importantly, the genetic and phenotypic traits possessed by late-stage amoeba-adapted P. aeruginosa are similar to what is observed for isolates obtained from chronic cystic fibrosis infections. This notable overlap in adaptation to different host types suggests similar selection pressures among host cell types. Author Summary Pseudomonas aeruginosa is an opportunistic pathogen that causes both acute infections in plants and animals, including humans and also causes chronic infections in immune compromised and cystic fibrosis patients. This bacterium is commonly found in soils and water where bacteria are constantly under threat of being consumed by the bacterial predators, protozoa. To escape being killed, bacteria have evolved a suite of mechanisms that protect them from being consumed or digested. Here we examined the effect of long-term predation on the genotype and phenotypes expressed by P. aeruginosa. We show that long-term co-incubation with protozoa resulted in mutations in the bacteria that made them less pathogenic. This is particularly interesting as we see similar mutations arise in bacteria associated with chronic infections. Thus, predation by protozoa and long term colonization of the human host may represent similar environments that select for similar losses in gene functions

Chemical interaction, interfacial effect and the microstructural characterization of the induced zinc–aluminum–Solanum tuberosum in chloride solution on mild steel

In this study, we report the effect of Solanum tuberosum (ST) as a strong additive on the morphological interaction, wear, and hardness properties of electroplated zinc coating in chloride bath solutions. The structural and the mechanical behavior of the Zn–Al–ST coating were studied and compared with the properties of Zn coatings. Characterization of the electrodeposited coatings were carried out using scanning electron microscopy, energy dispersive spectrometer, AFM, and X-ray diffraction techniques. The adhesion between the coatings and substrate was examined mechanically using hardness and wear techniques. From the results, amorphous Zn–Al–ST coatings were effectively obtained by electrodeposition using direct current. The coating morphology was revealed to be reliant on the bath composition containing strong leveling additives. From all indications, ST content contribute to a strong interfacial surface effect leading to crack-free and better morphology, good hardness properties, and improved wear resistance due to the precipitation of Zn2Si and Zn7Al2Si3. Hence, addition of ST is beneficial for the structural strengthening, hardness, and wear resistance properties of such coatings

Identification and structural analysis of C-terminally truncated collapsin response mediator protein-2 in a murine model of prion diseases

<p>Abstract</p> <p>Background</p> <p>Prion diseases are fatal neurodegenerative disorders that accompany an accumulation of the disease-associated form(s) of prion protein (PrP^Sc) in the central nervous system. The neuropathological changes in the brain begin with focal deposits of PrP^Sc, followed by pathomorphological abnormalities of axon terminal degeneration, synaptic loss, atrophy of dendritic trees, and eventual neuronal cell death in the lesions. However, the underlying molecular basis for these neuropathogenic abnormalities is not fully understood.</p> <p>Results</p> <p>In a proteomic analysis of soluble proteins in the brains of mice challenged intracerebrally with scrapie prion (Obihiro I strain), we found that the amount of the full-length form of collapsin response mediator protein-2 (CRMP-2; 61 kDa) decreased in the late stages of the disease, while the amount of its truncated form (56 kDa) increased to comparable levels observed for the full-length form. Detailed analysis by liquid chromatography-electrospray ionization-tandem mass spectrometry showed that the 56-kDa form (named CRMP-2-ΔC) lacked the sequence from serine⁵¹⁸to the C-terminus, including the C-terminal phosphorylation sites important for the regulation of axonal growth and axon-dendrite specification in developing neurons. The invariable size of the mRNA transcript in Northern blot analysis suggested that the truncation was due to post-translational proteolysis. By overexpression of CRMP-2-ΔC in primary cultured neurons, we observed the augmentation of the development of neurite branch tips to the same levels as for CRMP-2^T514A/T555A, a non-phosphorylated mimic of the full-length protein. This suggests that the increased level of CRMP-2-ΔC in the brain modulates the integrity of neurons, and may be involved in the pathogenesis of the neuronal abnormalities observed in the late stages of the disease.</p> <p>Conclusions</p> <p>We identified the presence of CRMP-2-ΔC in the brain of a murine model of prion disease. Of note, C-terminal truncations of CRMP-2 have been recently observed in models for neurodegenerative disorders such as ischemia, traumatic brain injury, and Wallerian degeneration. While the structural identity of CRMP-2-ΔC in those models remains unknown, the present study should provide clues to the molecular pathology of degenerating neurons in prion diseases in connection with other neurodegenerative disorders.</p

Not gods but animals : human dignity and vulnerable subjecthood

Drawing on earlier work on the conceptual structure of dignity, this paper will suggest a particular type of connectedness between vulnerability and human dignity; namely, that the ‘‘organizing idea’’ of human dignity is the idea of a particular sort of ethical response to universal human vulnerability. It is common ground among many, if not all, approaches to ethics that vulnerability requires us to respond ethically. Here, I argue that human dignity is distinctive among ethical values in that it values us because of, rather than in spite of, or regardless of, our universal vulnerability. The term ‘‘dignity’’ is used synonymously with ‘‘human dignity’’ here, since an investigation of the dignity of non-human entities forms no part of the present examination

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)1.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field

Cerebral microbleeds and stroke risk after ischaemic stroke or transient ischaemic attack: a pooled analysis of individual patient data from cohort studies

BACKGROUND: Cerebral microbleeds are a neuroimaging biomarker of stroke risk. A crucial clinical question is whether cerebral microbleeds indicate patients with recent ischaemic stroke or transient ischaemic attack in whom the rate of future intracranial haemorrhage is likely to exceed that of recurrent ischaemic stroke when treated with antithrombotic drugs. We therefore aimed to establish whether a large burden of cerebral microbleeds or particular anatomical patterns of cerebral microbleeds can identify ischaemic stroke or transient ischaemic attack patients at higher absolute risk of intracranial haemorrhage than ischaemic stroke. METHODS: We did a pooled analysis of individual patient data from cohort studies in adults with recent ischaemic stroke or transient ischaemic attack. Cohorts were eligible for inclusion if they prospectively recruited adult participants with ischaemic stroke or transient ischaemic attack; included at least 50 participants; collected data on stroke events over at least 3 months follow-up; used an appropriate MRI sequence that is sensitive to magnetic susceptibility; and documented the number and anatomical distribution of cerebral microbleeds reliably using consensus criteria and validated scales. Our prespecified primary outcomes were a composite of any symptomatic intracranial haemorrhage or ischaemic stroke, symptomatic intracranial haemorrhage, and symptomatic ischaemic stroke. We registered this study with the PROSPERO international prospective register of systematic reviews, number CRD42016036602. FINDINGS: Between Jan 1, 1996, and Dec 1, 2018, we identified 344 studies. After exclusions for ineligibility or declined requests for inclusion, 20 322 patients from 38 cohorts (over 35 225 patient-years of follow-up; median 1·34 years [IQR 0·19-2·44]) were included in our analyses. The adjusted hazard ratio [aHR] comparing patients with cerebral microbleeds to those without was 1·35 (95% CI 1·20-1·50) for the composite outcome of intracranial haemorrhage and ischaemic stroke; 2·45 (1·82-3·29) for intracranial haemorrhage and 1·23 (1·08-1·40) for ischaemic stroke. The aHR increased with increasing cerebral microbleed burden for intracranial haemorrhage but this effect was less marked for ischaemic stroke (for five or more cerebral microbleeds, aHR 4·55 [95% CI 3·08-6·72] for intracranial haemorrhage vs 1·47 [1·19-1·80] for ischaemic stroke; for ten or more cerebral microbleeds, aHR 5·52 [3·36-9·05] vs 1·43 [1·07-1·91]; and for ≥20 cerebral microbleeds, aHR 8·61 [4·69-15·81] vs 1·86 [1·23-1·82]). However, irrespective of cerebral microbleed anatomical distribution or burden, the rate of ischaemic stroke exceeded that of intracranial haemorrhage (for ten or more cerebral microbleeds, 64 ischaemic strokes [95% CI 48-84] per 1000 patient-years vs 27 intracranial haemorrhages [17-41] per 1000 patient-years; and for ≥20 cerebral microbleeds, 73 ischaemic strokes [46-108] per 1000 patient-years vs 39 intracranial haemorrhages [21-67] per 1000 patient-years). INTERPRETATION: In patients with recent ischaemic stroke or transient ischaemic attack, cerebral microbleeds are associated with a greater relative hazard (aHR) for subsequent intracranial haemorrhage than for ischaemic stroke, but the absolute risk of ischaemic stroke is higher than that of intracranial haemorrhage, regardless of cerebral microbleed presence, antomical distribution, or burden