PKCα translocation from mitochondria to nucleus is closely related to induction of apoptosis in gastric cancer cells

PKCs have been implicated in the regulation of cellular differentiation, proliferation, apoptosis and signal transduction. It was demonstrated in this study that PKCa was located both at mitochondria and in cytosol in gastric cancer cell line BGC-823. Treatment of cells with 12-O-tetradecanoylphorbol-13-acetate (TPA) resulted in the translocation of PKCa from both mitochondria and cytosol to nucleus as clearly shown by laser-scanning-confocal microscopy, while the protein level of PKCa was not changed by TPA treatment as detected by Western blot. The results also revealed that TPA-induced translocation of PKCa was in close association with apoptosis induction, and such association was further affirmed by other experiments where various apoptotic stimuli and specific inhibitors of PKC were used. Taken together, these findings indicate that translocation of PKCa from both mitochondria and cytosol to nucleus in gastric cancer cell is accompanied by induction of apoptosis, and may imply a new mechanism of th     

PKCα translocation from mitochondria to nucleus is closely related to induction of apoptosis in gastric cancer cells

PKCs have been implicated in the regulation of cellular differentiation, proliferation, apoptosis and signal transduction. It was demonstrated in this study that PKCα was located both at mitochondria and in cytosol in gastric cancer cell line BGC-823. Treatment of cells with 12-O-tetradecanoylphorbol-13-acetate (TPA) resulted in the translocation of PKCα from both mitochondria and cytosol to nucleus as clearly shown by laserscanningconfocal microscopy, while the protein level of PKCα was not changed by TPA treatment as detected by Western blot. The results also revealed that TPA-induced translocation of PKCα was in close association with apoptosis induction, and such association was further affirmed by other experiments where various apoptotic stimuli and specific inhibitors of PKC were used. Taken together, these findings indicate that translocation of PKCα from both mitochondria and cytosol to nucleus in gastric cancer cell is accompanied by induction of apoptosis, and may imply a new mechanism of the potential linking between cell apoptosis and PKCα translocation.     

Is MAC the knife that cuts cytochrome c from mitochondria during apoptosis?

Apoptosis is a phenomenon fundamental to higher eukaryotes and essential to mechanisms controlling tissue homeostasis. Bcl-2 family proteins tightly control this cell death program by regulating the permeabilization of the mitochondrial outer membrane and, hence, the release of cytochrome c and other proapoptotic factors. Mitochondrial apoptosis-induced channel (MAC) is the mitochondrial apoptosis-induced channel and is responsible for cytochrome c release early in apoptosis. MAC activity is detected by patch clamping mitochondria at the time of cytochrome c release. The Bcl-2 family proteins regulate apoptosis by controlling the formation of MAC. Depending on cell type and apoptotic inducer, Bax and/or Bak are structural component(s) of MAC. Overexpression of the antiapoptotic protein Bcl-2 eliminates MAC activity. The focus of this review is a biophysical characterization of MAC activity and its regulation by Bcl-2 family proteins, and ends with some discussion of therapeutic targets.     

Do urban areas act as foci for the spread of alien plant species? An assessment of temporal trends in the UK

Aim Given that urban landscapes often act as a point of entry for many non‐native species and urban development continues to increase as the human population rapidly expands, an understanding of the interaction between urbanization and non‐native plant species is important both in the control of potentially invasive species and in the conservation of native biodiversity. We investigated the spatial and temporal relationship between urban land cover and the distribution of non‐native species in Britain using two floristic data sets collected at two different time periods: 1987–88 and 2003–04. Location UK. Methods Using floristic data collected by the Botanical Society of the British Isles in 1987–88 (Monitoring Scheme) and 2003–04 (Local Change) in conjunction with habitat data obtained from the Land Cover Map of the UK, we conducted multiple regression analyses both within and between years on both groups of species (natives, neophytes and archaeophytes) and individual species. Results Neophytes (alien species introduced after 1500) were very strongly associated with urban land cover in both time periods and do not appear to be spreading out of urban habitats into the wider countryside. Archaeophytes (alien species introduced before 1500), however, showed a strong association with urban habitats in the earlier 1988 data set but no longer showed this association in the 2004 data set. Analysis at the individual species level showed that a large percentage of alien plant species, particularly archaeophytes, were not strongly associated with urban land cover or were negatively associated with such habitats. Main conclusions Our results suggest that there has been a reduction in the urban association of archaeophytes that is likely to have resulted from the recovery of archaeophytes associated with non‐urban (especially arable) habitats, following their decline in mid‐20th century, rather than from the movement of aliens into the wider countryside from urban habitats.     

Do environmental changes or juvenile competition act as mechanisms of species displacement in crayfishes?

The Big Creek Crayfish, Orconectes peruncus, is native to the St. Francis River drainage in Missouri, USA and is often absent where the introduced Woodland Crayfish, Orconectes hylas, has established. We performed a field experiment to determine whether effects of current abiotic conditions and interspecific competition with O. hylas were responsible for displacement of O. peruncus from parts of their former range. We examined growth and survival of juvenile male O. peruncus exposed to juvenile male O. hylas in enclosures at two sites in the former range of O. peruncus. Enclosures contained 8 (low density) or 16 individuals (high density) and had O. peruncus only (control) or both species (interspecific treatment). Juvenile O. peruncus were able to survive and grow in portions of their former range, implicating biotic versus abiotic factors in the displacement of O. peruncus. Survival rates of O. peruncus did not differ among treatments at either site. Orconectes peruncus showed significant growth in all treatments and interspecific effects were not greater than intraspecific effects on O. peruncus growth rates. High-density treatments showed significantly reduced O. peruncus growth rates compared to low-density treatments, except in Carver Creek interspecific treatments. When considered in the context of previous studies examining the effects of O. hylas on O. peruncus, results suggest that neither direct competition between juvenile males of the two species or abiotic change are responsible for the decreased range of O. peruncus. Additional research is required to determine the mechanism(s) driving the displacement of O. peruncus.     

Do modulators of the mitochondrial K(ATP) channel change the function of mitochondria in situ?

Pharmacological opening of mitochondrial cardiac ATP-sensitive potassium (K(ATP)) channels has the chance to be a promising but still controversial cardioprotective mechanism. Physiological roles of mitochondrial K(ATP) channels in the myocardium remain unclear. We studied the effects of diazoxide, a specific opener of these channels, on the function of rat mitochondria in situ in saponin-permeabilized fibers using an ionic medium that mimics the cytosol. In the presence of NADH-producing substrates (malate + glutamate), neither 100 microm diazoxide nor 100 microm glibenclamide (a K(ATP) channel blocker) changed the mitochondrial respiration in the absence or presence of ADP. Because the K(ATP) channel function could be modified by changes in adenine nucleotide concentrations near the mitochondria, we studied the effects of diazoxide and glibenclamide on the functional activity of mitochondrial kinases. Both diazoxide and glibenclamide did not change the in situ ADP sensitivity in the presence or absence of creatine (apparent K(m) values for ADP were, respectively, 59 +/- 9 and 379 +/- 45 microm). Similarly, stimulation of the mitochondrial respiration with AMP in the presence of ATP due to adenylate kinase activity was not affected by the modulators of K(ATP) channels. However, when succinate was used as substrate, diazoxide significantly inhibited basal respiration by 22% and maximal respiration by 24%. Thus, at a cardioprotective dose, the main functional effect of diazoxide depends on respiratory substrates and seems not to be related to K(ATP) channel activity.     

The GABAergic network in the suprachiasmatic nucleus as a key regulator of the biological clock: does it change during senescence?

GABA is the main neurotransmitter of the hypothalamic suprachiasmatic nucleus (SCN) and plays a key role in the function of this master circadian pacemaker. Despite the evidence that disturbances of biological rhythms are common during aging, little is known about the GABAergic network in the SCN of the aging brain. We here provide a brief overview of the GABAergic structures and the role of GABA in the SCN. We also review some age-related changes of the GABAergic system occurring in the brain outside the SCN. Finally, we present preliminary data on the GABAergic system within the SCN comparing young and aging mice. In particular, our study on age-related changes in the SCN focused on the daily expression of the alpha3 subunit of the GABA(A) receptor and on the density of GABAergic axon terminals. Interestingly, our preliminary findings point to alterations of the GABAergic network in the biological clock during senescence.     

The GABAergic network in the suprachiasmatic nucleus as a key regulator of the biological clock: does it change during senescence?

GABA is the main neurotransmitter of the hypothalamic suprachiasmatic nucleus (SCN) and plays a key role in the function of this master circadian pacemaker. Despite the evidence that disturbances of biological rhythms are common during aging, little is known about the GABAergic network in the SCN of the aging brain. We here provide a brief overview of the GABAergic structures and the role of GABA in the SCN. We also review some age‐related changes of the GABAergic system occurring in the brain outside the SCN. Finally, we present preliminary data on the GABAergic system within the SCN comparing young and aging mice. In particular, our study on age‐related changes in the SCN focused on the daily expression of the α3 subunit of the GABA_A receptor and on the density of GABAergic axon terminals. Interestingly, our preliminary findings point to alterations of the GABAergic network in the biological clock during senescence.     

Do the oscillations of cardiovascular parameters persist during voluntary apnea in humans?

The aim of this study was to ascertain the persistence of heart rate and blood pressure oscillations at the onset of voluntary apnea in humans and to assess the dependence of the fluctuations parameters on the chemoreceptor activity. In 24 young subjects (10 males, 14 females, mean age 20.4 years) heart rate (represented by its reciprocal value--RR-intervals), systolic blood pressure (SBP) and diastolic blood pressure (DBP) during controlled breathing (CB) of atmospheric air and oxygen followed by apnea were recorded continuously. The cosine functions were then fitted by nonlinear regression analysis to the heart rate, SBP and DBP oscillations during CB and at the onset of apnea. The parameters of oscillations were different during atmospheric air breathing compared to oxygen breathing. During oxygen breathing there was an increase of the RR-interval oscillations--relative bradycardia and enhanced magnitude of respiratory sinus arythmia. During apnea, the base level of the blood pressure oscillations was higher after breathing of atmospheric air compared to oxygen breathing. At least one cosine-like wave oscillation was present at the onset of apnea in the heart rate, SBP and DBP and the second wave was present in all assessed parameters in at least 70% of recordings. The oscillations in RR-intervals are, to some extent, independent of blood pressure oscillations. No significant gender differences were found either in the duration of breath holding or in the RR and SBP oscillations parameters.     

Calcium-sensing receptor activating phosphorylation of PKC�� translocation on mitochondria to induce cardiomyocyte apoptosis during ischemia/reperfusion

The calcium-sensing receptor (CaSR) is a G protein-coupled receptor (GPCR) that activates intracellular effectors; for example, it causes inositol phosphate (IP) and 1,2 diacylglycerol (DAG) accumulation, stimulating the release of intracellular calcium and the activation of the protein kinase Cs (PKCs). The activation of CaSR by ischemia/reperfusion (I/R) induces cardiomyocyte apoptosis through the mitochondrial apoptotic pathway; however, the underlying mechanisms remain unclear. In this study, rat hearts were subjected to 30 min of ischemia followed by 2 h of reperfusion in the presence of a CaSR activator, GdCl₃. Our results revealed that, under these conditions, the expression of CaSR was increased, the number of apoptotic cardiomyocytes was significantly increased (as shown by terminal deoxy-nucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay) and the cells with a typical apoptotic morphology were observed using transmission electron microscopy. Our data further showed that mitochondria isolated from hearts that had undergone I/R combined with GdCl₃ exhibited a significant increase in the translocation of phosphorylated PKC?? to the mitochondria, an increase in cytochrome c (cyt c) release from the mitochondria and a marked decrease in mitochondrial potential. The administration of rottlerin, an inhibitor of PKC??, significantly reduced reperfusion-induced apoptosis, phospho-PKC?? translocation to the mitochondria and the release of cyt c from the mitochondria. Thus, the involvement of CaSR in cardiac apoptosis through the mitochondrial pathway during I/R with GdCl₃ is related to phospho-PKC?? translocation to the mitochondria.     

Does the redox status of cytochrome C act as a fail-safe mechanism in the regulation of programmed cell death?

It has now become recognized that one of the key events in the induction of apoptosis, or programmed cell death, in both plants and animals is the release of cytochrome c from mitochondria. It is also known that oxidative stress imposed on cells can have a profound effect on the onset or progression of apoptosis. Here, we discuss how the redox status of cytochrome c, and thus its structure, can be altered by the presence of reactive oxygen species (ROS) and reduced glutathione (GSH). We suggest that cytochrome c will only induce programmed cell death if present in the cytoplasm in the oxidized state, and that the presence of high levels of cytoplasmic GSH maintain cytochrome c in an inactive (reduced) state, thus behaving as a fail-safe mechanism if cytochrome c is released by mitochondria when programmed cell death is not the required outcome. If the redox status of the cell is disturbed however, perhaps in the presence of hydrogen peroxide, GSH concentrations will drop, the cellular E(h) will rise, and cytochrome c will tend towards the oxidized state, allowing programmed cell death to proceed. Therefore, we propose that the redox state of cytoplasmic cytochrome c may be a key regulator of programmed cell death.     

Does the respiratory rate in sea urchin embryos increase during early development without proliferation of mitochondria?

During early development of the sea urchin, the respiratory rate, enhanced upon fertilization, is maintained up to hatching (pre-hatching period) and then gradually increases to a maximum at the gastrula stage (post-gastrula period). Except for a short duration after fertilization, respiration in embryos is strongly inhibited by CN⁻ and antimycin A. During the whole span of early development, the amounts of proteins, cytochromes and the specific activities of cytochrome c oxidase and reduced nicotinamide adenine dinucleotide (NADH) cytochrome c reductase in mitochondria are practically the same as in unfertilized eggs. A marked augmentation of mitochondrial respiration after hatching probably occurs without net increase in whole mitochondrial intrinsic capacities. Carbonylcyanide p-trifluoromethoxyphenylhydrazone (FCCP) or tetramethyl p-phenylenediamine (TMPD) enhances the respiratory rate in the pre-hatching period but hardly augments the respiration in the post-gastrula period. In the presence of both FCCP and TMPD, the respiratory rate in the pre-hatching period was as high as in the post-gastrula period. Probably, electron transport in the mitochondrial respiratory chain is regulated by acceptor control and limitation of cytochrome c reduction in the pre-hatching period and released from those regulations in the post-gastrula period. Acceptor control of respiration is experimentally reproduced in isolated mitochondria by making adenine nucleotide levels as those levels in the pre-hatching period.     

How Do Tracking and Changes in Dietary Pattern during Adolescence Relate to the Amount of Body Fat in Early Adulthood?

Background

Few studies have addressed the influence of dietary patterns (DP) during adolescence on the amount of body fat in early adulthood.

Objective

To analyze the associations between DP tracking and changes in the period between 15 and 18 years of age and the percentage of body fat (%BF) at age 18 years.

Methods

We used data from 3,823 members of the 1993 Pelotas (Brazil) birth cohort. Body density was measured at age 18 years by air displacement plethysmograph (BOD POD) and the %BF was calculated applying the Siri equation. Based on the estimates from the FFQ, we identified DP at ages 15 (“Varied”, “Traditional”, “Dieting” and “Processed meats”) and 18 years (“Varied”, “Traditional”, “Dieting” and “Fish, fast food and alcohol”). The DP tracking was defined as the individual’s adherence to the same DP at both ages. Associations were tested using multiple linear regression models stratified by sex.

Results

The mean %BF was 25.0% (95% CI: 24.7 to 25.4), significantly greater for girls than boys (p<0.001). The adherence to any DP at age 15 years was not associated with the %BF at age 18 years. However, individuals who adhered to a “Dieting” DP at age 18 years showed greater %BF (1.30 and 1.91 percentage points in boys and girls, respectively) in comparison with those who adhered to a “Varied” DP. Boys who presented tracking of a “Dieting” DP presented greater average %BF in comparison with others DP, as well as girls who changed from the “Traditional” or “Processed meats” DP to a “Dieting” DP.

Conclusion

These results may support public health policies and strategies focused on improving dietary habits of adolescents and young adults and preventing accumulation of body fat, especially among the adolescents with restrictive dietary habits.     

Prolines in βA-sheet of neural cadherin act as a switch to control the dynamics of the equilibrium between monomer and dimer

Neural cadherins dimerize through the formation of calcium-dependent strand-crossover structures. Dimerization of cadherins leads to cell-cell adhesion in multicellular organisms. Strand-crossover dimer forms exclusively between the first N-terminal extracellular modules (EC1) of the adhesive partners via swapping of their βA-sheets and docking of tryptophan-2 in the hydrophobic pocket. In the apo-state wild-type cadherin is predominantly monomer, which indicates that the dimerization is energetically unfavorable in the absence of calcium. Addition of calcium favors dimer formation by creating strain in the monomer and lowering the energetic barrier between monomer and dimer. Dynamics of the monomer-dimer equilibrium is vital for plasticity of synapses. Prolines recurrently occur in proteins that form strand-crossover dimer and are believed to be the source of the strain in the monomer. N-cadherins have two proline residues in the βA-sheet. We focused our studies on the role of these two prolines in calcium-dependent dimerization. Spectroscopic, electrophoretic, and chromatopgraphic studies showed that mutations of both prolines to alanines increased the dimerization affinity by ~20-fold and relieved the requirement of calcium in dimerization. The P5A and P6A mutant formed very stable dimers that required denaturation of protein to disassemble in the apo conditions. In summary, the proline residues act as a switch to control the dynamics of the equilibrium between monomer and dimer which is crucial for the plasticity of synapses.