The ceramide kinase inhibitor NVP-231 inhibits breast and lung cancer cell proliferation by inducing M phase arrest and subsequent cell death

Background and Purpose Ceramide kinase (CerK) catalyzes the generation of ceramide-1-phosphate which may regulate various cellular functions, including inflammatory reactions and cell growth. Here, we studied the effect of a recently developed CerK inhibitor, NVP-231, on cancer cell proliferation and viability and investigated the role of cell cycle regulators implicated in these responses. Experimental Approach The breast and lung cancer cell lines MCF-7 and NCI-H358 were treated with increasing concentrations of NVP-231 and DNA synthesis, colony formation and cell death were determined. Flow cytometry was performed to analyse cell cycle distribution of cells and Western blot analysis was used to detect changes in cell cycle regulator expression and activation. Key Results In both cell lines, NVP-231 concentration-dependently reduced cell viability, DNA synthesis and colony formation. Moreover it induced apoptosis, as measured by increased DNA fragmentation and caspase-3 and caspase-9 cleavage. Cell cycle analysis revealed that NVP-231 decreased the number of cells in S phase and induced M phase arrest with an increased mitotic index, as determined by increased histone H3 phosphorylation. The effect on the cell cycle was even more pronounced when NVP-231 treatment was combined with staurosporine. Finally, overexpression of CerK protected, whereas down-regulation of CerK with siRNA sensitized, cells for staurosporine-induced apoptosis. Conclusions and Implications Our data demonstrate for the first time a crucial role for CerK in the M phase control in cancer cells and suggest its targeted inhibition, using drugs such as NVP-231, in combination with conventional pro-apoptotic chemotherapy.

Physical activity in schizophrenia is higher in the first episode than in subsequent ones

Schizophrenia is frequently associated with abnormal motor behavior, particularly hypokinesia. The course of the illness tends to deteriorate in the first years. We aimed to assess gross motor activity in patients with a first episode (n = 33) and multiple episodes (n = 115) of schizophrenia spectrum disorders using wrist actigraphy. First episode patients were younger, had higher motor activity and reduced negative symptom severity. Covarying for age, chlorpromazine equivalents, and negative symptoms, first episode patients still had higher motor activity. This was also true after excluding patients with schizophreniform disorder from the analyses. In first episode patients, but not in patients with multiple episodes, motor activity was correlated with antipsychotic dosage. In conclusion, after controlling for variables related to disorder chronicity, patients with first episodes were still more active than patients with multiple episodes. Thus, reduced motor activity is a marker of deterioration in the course of schizophrenia spectrum disorders.

More conflict does not trigger more adjustment of cognitive control for subsequent performance: A study of the bivalency effect

Encountering a conflict triggers an adjustment of cognitive control. This adjustment of cognitive control can even affect subsequent performance. The purpose of the present study was to determine whether more conflict triggers more adjustment of cognitive control for subsequent performance. To this end, we focussed on the bivalency effect, that is, the adjustment of cognitive control following the conflict induced by bivalent stimuli (i.e., stimuli with relevant features for two tasks). In two experiments, we tested whether the amount of conflict triggered by bivalent stimuli affected the bivalency effect. Bivalent stimuli were either compatible (i.e., affording one response) or incompatible (i.e., affording two different responses). Thus, compatible bivalent stimuli involved a task conflict, whereas incompatible bivalent stimuli involved a task and a response conflict. The results showed that the bivalency effect was not affected by this manipulation. This indicates that more conflict does not trigger more adjustment of cognitive control for subsequent performance. Therefore, only the occurrence of conflict – not its amount – is determinant for cognitive control

Role of a ribosomal RNA phosphate oxygen during the EF-G–triggered GTP hydrolysis

Elongation factor-catalyzed GTP hydrolysis is a key reaction during the ribosomal elongation cycle. Recent crystal structures of G proteins, such as elongation factor G (EF-G) bound to the ribosome, as well as many biochemical studies, provide evidence that the direct interaction of translational GTPases (trGTPases) with the sarcin-ricin loop (SRL) of ribosomal RNA (rRNA) is pivotal for hydrolysis. However, the precise mechanism remains elusive and is intensively debated. Based on the close proximity of the phosphate oxygen of A2662 of the SRL to the supposedly catalytic histidine of EF-G (His87), we probed this interaction by an atomic mutagenesis approach. We individually replaced either of the two nonbridging phosphate oxygens at A2662 with a methyl group by the introduction of a methylphosphonate instead of the natural phosphate in fully functional, reconstituted bacterial ribosomes. Our major finding was that only one of the two resulting diastereomers, the SP methylphosphonate, was compatible with efficient GTPase activation on EF-G. The same trend was observed for a second trGTPase, namely EF4 (LepA). In addition, we provide evidence that the negative charge of the A2662 phosphate group must be retained for uncompromised activity in GTP hydrolysis. In summary, our data strongly corroborate that the nonbridging proSP phosphate oxygen at the A2662 of the SRL is critically involved in the activation of GTP hydrolysis. A mechanistic scenario is supported in which positioning of the catalytically active, protonated His87 through electrostatic interactions with the A2662 phosphate group and H-bond networks are key features of ribosome-triggered activation of trGTPases.

Distribution of cutin and suberin biomarkers under forest trees with different root systems

Background and aims Differences in chemical composition of root compounds and root systems among tree species may affect organic matter (OM) distribution, source and composition in forest soils. The objective of this study was to elucidate the contribution of species specific cutin and suberin biomarkers as proxies for shoot- and root-derived organic carbon (OC) to soil OM at different depths with increasing distance to the stems of four different tree species. Methods The contribution of cutin- and suberin-derived lipids to OM in a Cutanic Alisol was analyzed with increasing soil depth and distance to the stems of Fagus sylvatica L., Picea abies (L.) Karst., Quercus robur L. and Pseudotsuga menziesii (Mirb.) Franco. Cutin and suberin monomers of plants and soils were analyzed by alkaline hydrolysis and subsequent gas chromatography–mass spectrometry. Results The amount and distribution of suberin-derived lipids in soil clearly reflected the specific root system of the different tree species. The amount of cutin-derived lipids decreased strongly with soil depth, indicating that the input of leaf/needle material is restricted to the topsoil. In contrast to the suberin-derived lipids, the spatial pattern of cutin monomer contribution to soil OM did not depend on tree species. Conclusions Our results document the importance of tree species as a main factor controlling the composition and distribution of OM in forest soils. They reveal the impact of tree species on root-derived OM distribution and the necessity to distinguish among different zones when studying soil OM storage in forests.

Role of a ribosomal RNA phosphate oxygen during the EF-G-triggered hydrolysis

Elongation factor-catalyzed GTP hydrolysis is a key reaction during the ribosomal elongation cycle. Recent crystal structures of G proteins, such as elongation factor G (EF-G) bound to the ribosome, as well as many biochemical studies, provide evidence that the direct interaction of translational GTPases (trGTPases) with the sarcin-ricin loop (SRL) of ribosomal RNA (rRNA) is pivotal for hydrolysis. However, the precise mechanism remains elusive and is intensively debated. Based on the close proximity of the phosphate oxygen of A2662 of the SRL to the supposedly catalytic histidine of EF-G (His87), we probed this interaction by an atomic mutagenesis approach. We individually replaced either of the two nonbridging phosphate oxygens at A2662 with a methyl group by the introduction of a methylphosphonate instead of the natural phosphate in fully functional, reconstituted bacterial ribosomes. Our major finding was that only one of the two resulting diastereomers, the SP methylphosphonate, was compatible with efficient GTPase activation on EF-G. The same trend was observed for a second trGTPase, namely EF4 (LepA). In addition, we provide evidence that the negative charge of the A2662 phosphate group must be retained for uncompromised activity in GTP hydrolysis. (1) In summary, our data strongly corroborate that the nonbridging proSP phosphate oxygen at the A2662 of the SRL is critically involved in the activation of GTP hydrolysis. A mechanistic scenario is supported in which positioning of the catalytically active, protonated His87 through electrostatic interactions with the A2662 phosphate group and H-bond networks are key features of ribosome-triggered activation of trGTPases.

Role of a ribosomal RNA phosphate oxygen during the EF-G-triggered hydrolysis

Elongation factor-catalyzed GTP hydrolysis is a key reaction during the ribosomal elongation cycle. Recent crystal structures of G proteins, such as elongation factor G (EF-G) bound to the ribosome, as well as many biochemical studies, provide evidence that the direct interaction of translational GTPases (trGTPases) with the sarcin-ricin loop (SRL) of ribosomal RNA (rRNA) is pivotal for hydrolysis. However, the precise mechanism remains elusive and is intensively debated. Based on the close proximity of the phosphate oxygen of A2662 of the SRL to the supposedly catalytic histidine of EF-G (His87), we probed this interaction by an atomic mutagenesis approach. We individually replaced either of the two nonbridging phosphate oxygens at A2662 with a methyl group by the introduction of a methylphosphonate instead of the natural phosphate in fully functional, reconstituted bacterial ribosomes. Our major finding was that only one of the two resulting diastereomers, the SP methylphosphonate, was compatible with efficient GTPase activation on EF-G. The same trend was observed for a second trGTPase, namely EF4 (LepA). In addition, we provide evidence that the negative charge of the A2662 phosphate group must be retained for uncompromised activity in GTP hydrolysis. (1) In summary, our data strongly corroborate that the nonbridging proSP phosphate oxygen at the A2662 of the SRL is critically involved in the activation of GTP hydrolysis. A mechanistic scenario is supported in which positioning of the catalytically active, protonated His87 through electrostatic interactions with the A2662 phosphate group and H-bond networks are key features of ribosome-triggered activation of trGTPases.

Protein Hydrolysis and Nitrogen Remobilisation in Plant Life and Senescence

In plant cells, as in all other cells, proteins are submitted to permanent turnover, and the intracellular content of a given protein depends on its rate of both synthesis and degradation. The life time of most proteins is shorter than that of the cell. Thus, in young leaves of Lemna minor, the average half-life of protein was estimated to be 7 days, and it was shorter under stress conditions (Davies 1982). Such observations mean that nitrogen and amino acid fluxes are both cylic and permanent. Although protein turnover may appear wasteful, in terms of energy, numerous studies have shown that proteolysis provides multiple functions in cell physiology, and is an essential regulatory mechanism of cell metabolism and development.

Prospective longitudinal study of subcortical brain volumes in individuals at high familial risk of mood disorders with or without subsequent onset of depression

Subcortical volumetric brain abnormalities have been observed in mood disorders. However, it is unknown whether these reflect adverse effects predisposing to mood disorders or emerge at illness onset. Magnetic resonance imaging was conducted at baseline and after two years in 111 initially unaffected young adults at increased risk of mood disorders because of a close family history of bipolar disorder and 93 healthy controls (HC). During the follow-up, 20 high-risk subjects developed major depressive disorder (HR-MDD), with the others remaining well (HR-well). Volumes of the lateral ventricles, caudate, putamen, pallidum, thalamus, hippocampus and amygdala were extracted for each hemisphere. Using linear mixed-effects models, differences and longitudinal changes in subcortical volumes were investigated between groups (HC, HR-MDD, HR-well). There were no significant differences for any subcortical volume between groups controlling for multiple testing. Additionally, no significant differences emerged between groups over time. Our results indicate that volumetric subcortical brain abnormalities of these regions using the current method appear not to form familial trait markers for vulnerability to mood disorders in close relatives of bipolar disorder patients over the two-year time period studied. Moreover, they do not appear to reduce in response to illness onset at least for the time period studied.