STRUCTURE,LIFE HISTORY AND SYSTEMATICS OF RHOICOSPHENIA (BACILLARIOPHYTA). IV. CORRELATION OF SIZE REDUCTION WITH CHANGES IN VALVE MORPHOLOGY OF RH. GENUFLEXA1

Rhoicosphenia Grun. is a relatively isolated genus among the biraphid diatoms. Morphological changes in an isopolar member of the genus, Rh. genuflexa (Kütz.) Medlin, were investigated using light and scanning electron microscopy. The fully raphid valve showed changes in its flexure that could be correlated with size reduction during its life history from the initial cells to the smallest cells found in the population. Bands showed changes in number (from three to one) related to size reduction. Rh. genuflexa is morphologically similar to Rh. abbreviata (C. Ag.) Lange-Bert. (=Rh. curvata (Kütz.) Grun.), although the two are distinct taxa. These observations support previous contentions that Rhoicosphenia is a natural taxonomic grouping.     

Growth and survival of Daphnia in epilimnetic and metalimnetic water from oligotrophic lakes: the effects of food and temperature

SUMMARY 1. In oligotrophic lakes, phytoplankton and bacteria growing in the deep chlorophyll maximum in the cool metalimnion of lakes often dominate biomass and production, but the importance of this source of food for zooplankton is unknown.
2. During much of the day, Daphnia rosea in two mountain lakes inhabited deep chlorophyll layers where food availability was at least equal to that in the epilimnion.
3. To determine the importance of the two strata (epilimnion and metalimnion) for Daphnia , we used a cross-classified factorial experiment to measure how epilimnetic and metalimnetic food and temperature (10 and 16 °C) influenced survival, growth and reproduction.
4. Daphnia survived and grew better when fed seston from the epilimnion of one lake, although chlorophyll, particulate nitrogen and particulate carbon were 2–2.5 times greater in the metalimnion.
5. Temperature had no significant influence on Daphnia survival or growth. Similar results were obtained with food from the second lake, with Daphnia surviving and reproducing better when provided with epilimnetic, rather than metalimnetic food, although the quantities of chlorophyll and carbon in the two strata were similar.
6. Food quality, rather than quantity or temperature, appeared to be the most important determinant influencing survival, growth and reproduction, and the greater food quantity in the metalimnia was not used effectively by the Daphnia .     

Detection of apolipoprotein B100 early conformational changes during oxidation

Conformational changes of human plasma apolipoprotein B100 (apoB) during oxidative modification of low-density lipoproteins (LDL) have been investigated. Emphasis has been put on the early stages of LDL oxidation and the modification of apoB. We have applied two different modes of LDL oxidation initiation in order to approach the problem from different perspectives. To study conformational changes of the protein and the phospholipids surface monolayer, we have applied attenuated total reflection infrared as well as fluorescence spectroscopy. We have found for the first time that conformational changes of apoB occur even in the earliest stages of oxidation process and that those are located predominantly in the beta-sheet regions. The dynamics of changes has also been described and related to different stages of oxidation. After initial increase in particle surface accessibility and mobility, by entering into the propagation phase of oxidation process, LDL surface accessibility and mobility are decreased. Finally, in the decomposition phase of LDL oxidation, as the particle faces large chemical and physical changes, surface mobility and accessibility is increased again. These observations provide new insights into the modifications of LDL particles upon oxidation.     

Adhesion to medical device materials and biofilm formation capability of some species of enterococci in different physiological states

Enterococci may survive in adverse environments including the human body where bacteriocins, antibiotics, iron-limitation and immune response represent stressing conditions for bacteria that cause division block. In those conditions, bacteria present in the human body would hardly be in an exponentially growing phase but would mostly be in physiological states such as starvation or the viable but nonculturable (VBNC) state. The possibility that the starved and VBNC bacteria can maintain their ability to adhere to living and inanimate substrates is the first mandatory step for them potentially to cause an infection process. In this study it is shown that starved and stationary enterococcal cells are able to form biofilms on plastic material albeit with reduced efficiency as compared to growing cells. Moreover, although VBNC enterococcal forms are not capable of forming biofilms, Enterococcus faecalis and other enterococcal species of medical interest maintain their ability to synthesize the polymeric matrix for a limited period of time under adverse environmental conditions. The data presented, together with those regarding the maintenance of the division recovery potential already proved in nonculturable bacteria, further support the possibility for the VBNC and other nondividing bacterial forms to have a role as infectious agents and to constitute a risk to human health.     

Feed‐backs among inbreeding,inbreeding depression in sperm traits,and sperm competition can drive evolution of costly polyandry

Ongoing ambitions are to understand the evolution of costly polyandry and its consequences for species ecology and evolution. Emerging patterns could stem from feed‐back dynamics between the evolving mating system and its genetic environment, defined by interactions among kin including inbreeding. However, such feed‐backs are rarely considered in nonselfing systems. We use a genetically explicit model to demonstrate a mechanism by which inbreeding depression can select for polyandry to mitigate the negative consequences of mating with inbred males, rather than to avoid inbreeding, and to elucidate underlying feed‐backs. Specifically, given inbreeding depression in sperm traits, costly polyandry evolved to ensure female fertility, without requiring explicit inbreeding avoidance. Resulting sperm competition caused evolution of sperm traits and further mitigated the negative effect of inbreeding depression on female fertility. The evolving mating system fed back to decrease population‐wide homozygosity, and hence inbreeding. However, the net overall decrease was small due to compound effects on the variances in sex‐specific reproductive success and paternity skew. Purging of deleterious mutations did not eliminate inbreeding depression in sperm traits or hence selection for polyandry. Overall, our model illustrates that polyandry evolution, both directly and through sperm competition, might facilitate evolutionary rescue for populations experiencing sudden increases in inbreeding.     

Colocalization of cellular nanostructure using confocal fluorescence and partial wave spectroscopy

A new multimodal confocal microscope has been developed, which includes a parallel Partial Wave Spectroscopic (PWS) microscopy path. This combination of modalities allows molecular‐specific sensing of nanoscale intracellular structure using fluorescent labels. Combining molecular specificity and sensitivity to nanoscale structure allows localization of nanostructural intracellular changes, which is critical for understanding the mechanisms of diseases such as cancer. To demonstrate the capabilities of this multimodal instrument, we imaged HeLa cells treated with valinomycin, a potassium ionophore that uncouples oxidative phosphorylation. Colocalization of fluorescence images of the nuclei (Hoechst 33342) and mitochondria (anti‐mitochondria conjugated to Alexa Fluor 488) with PWS measurements allowed us to detect a significant decrease in nuclear nanoscale heterogeneity (Σ), while no significant change in Σ was observed at mitochondrial sites. In addition, application of the new multimodal imaging approach was demonstrated on human buccal samples prepared using a cancer screening protocol. These images demonstrate that nanoscale intracellular structure can be studied in healthy and diseased cells at molecular‐specific sites.

     

Focal adhesion kinase plays a dual role in TRAIL resistance and metastatic outgrowth of malignant melanoma

Despite remarkable advances in therapeutic interventions, malignant melanoma (MM) remains a life-threating disease. Following high initial response rates to targeted kinase-inhibition metastases quickly acquire resistance and present with enhanced tumor progression and invasion, demanding alternative treatment options. We show 2^nd generation hexameric TRAIL-receptor-agonist IZI1551 (IZI) to effectively induce apoptosis in MM cells irrespective of the intrinsic BRAF/NRAS mutation status. Conditioning to the EC₅₀ dose of IZI converted the phenotype of IZI-sensitive parental MM cells into a fast proliferating and invasive, IZI-resistant metastasis. Mechanistically, we identified focal adhesion kinase (FAK) to play a dual role in phenotype-switching. In the cytosol, activated FAK triggers survival pathways in a PI3K- and MAPK-dependent manner. In the nucleus, the FERM domain of FAK prevents activation of wtp53, as being expressed in the majority of MM, and consequently intrinsic apoptosis. Caspase-8-mediated cleavage of FAK as well as FAK knockdown, and pharmacological inhibition, respectively, reverted the metastatic phenotype-switch and restored IZI responsiveness. FAK inhibition also re-sensitized MM cells isolated from patient metastasis that had relapsed from targeted kinase inhibition to cell death, irrespective of the intrinsic BRAF/NRAS mutation status. Hence, FAK-inhibition alone or in combination with 2nd generation TRAIL-receptor agonists may be recommended for treatment of initially resistant and relapsed MM, respectively.Subject terms: Metastasis, Target identification     

Human haptoglobin structure and function--a molecular modelling study

Hemoglobin is the most prominent protein in blood, transporting O(2) and facilitating reactive oxygen and nitrogen species detoxification. Hemoglobin metabolism leads to the release of extra-erythrocytic hemoglobin, with potentially severe consequences for health. Extra-erythrocytic hemoglobin is complexed to haptoglobin for clearance by tissue macrophages. The human gene for haptoglobin consists of three structural alleles: Hp1F, Hp1S and Hp2. The products of the Hp1F and Hp1S alleles differ by only one amino acid, whereas the Hp2 allele is the result of a fusion of the Hp1F and Hp1S alleles, is present only in humans and gives rise to a longer alpha-chain. Haptoglobin consists of a dimer of alphabeta-chains covalently linked by a disulphide bond between the Cys15 residue of each alpha-chain. However, the presence of the Hp1 and Hp2 alleles in humans gives rise to HPT1-1 dimers (covalently linked by Cys15 residues), HPT1-2 hetero-oligomers and HPT2-2 oligomers. In fact, the HPT2 variant displays two free Cys residues (Cys15 and Cys74) whose participation in intermolecular disulphide bonds gives rise to higher-order covalent multimers. Here, the complete modelling of both haptoglobin variants, together with their basic quaternary structure arrangements (i.e. HPT1 dimer and HPT2 trimer), is reported. The structural details of the models, which represent the first complete view of the molecular details of human haptoglobin variants, are discussed in relation to the known haptoglobin function(s).     

Modulation of the inward rectifier potassium channel IRK1 by the Ras signaling pathway.

In this study, we investigated the role of Ras and the mitogen-activated protein kinase (MAPK) pathway in the modulation of the inward rectifier potassium channel IRK1. We show that although expression of IRK1 in HEK 293 cells leads to the appearance of a potassium current with strong inward rectifying properties, coexpression of the constitutively active form of Ras (Ras-L61) results in a significant reduction of the mean current density without altering the biophysical properties of the channel. The inhibitory effect of Ras-L61 is not due to a decreased expression of IRK1 since Northern analysis indicates that IRK1 mRNA level is not affected by Ras-L61 co-expression. Moreover, the inhibition can be relieved by treatment with the mitogen-activated protein kinase/ERK kinase (MEK) inhibitor PD98059. Confocal microscopy analysis of cells transfected with the fusion construct green fluorescent protein-IRK1 shows that the channel is mainly localized at the plasma membrane. Coexpression of Ras-L61 delocalizes fluorescence to the cytoplasm, whereas treatment with PD98059 partially restores the membrane localization. In conclusion, our data indicate that the Ras-MAPK pathway modulates IRK1 current by affecting the subcellular localization of the channel. This suggests a role for Ras signaling in regulating the intracellular trafficking of this channel.