The inner nuclear membrane protein lamin B receptor forms distinct microdomains and links epigenetically marked chromatin to the nuclear envelope

Using heterochromatin-enriched fractions, we have detected specific binding of mononucleosomes to the N-terminal domain of the inner nuclear membrane protein lamin B receptor. Mass spectrometric analysis reveals that LBR-associated particles contain complex patterns of methylated/acetylated histones and are devoid of "euchromatic" epigenetic marks. LBR binds heterochromatin as a higher oligomer and forms distinct nuclear envelope microdomains in vivo. The organization of these membrane assemblies is affected significantly in heterozygous ic (ichthyosis) mutants, resulting in a variety of structural abnormalities and nuclear defects.     

Heat-induced degradation of overexpressed glucocorticoid receptor Separate protective roles of hsp90 and hsp70

The glucocorticoid receptor (GR) occurs in cells in the form of a hormone-responsive complex (HRC) with hsp90. The HRC is dynamic, with hsp90 constantly directing disassembly, and hsp70, assisted by hsp90, driving reassembly. WCL2 cells stably overexpress GR to an extent that reduces the excess of hsp90 and hsp70 over GR by about 10-fold, compared to the ratio in HeLa cells. Yet the half-lives of the HRC in WCL2 and HeLa cells are comparable. As a result, the rate of assembly in WCL2 is overwhelmed by accumulation of the non-hormone-binding form of GR in its complex with hsp70 and hsp90. This form comprised some 50% of total GR in WCL2 cells. When the cells were heated to 44 degrees C, the hormone-binding activity and solubility of GR fell in parallel, and the receptor formed heavy aggregates by sequestering large amounts of hsp70. About 40% of this aggregated receptor was degraded in cells recovering at 37 degrees C in the presence of cycloheximide. Concentration of GR protein increased with increasing induction of hsp70 following exposure to 41-44 degrees C. However, balance between hormone-binding and inert forms of GR could shift in either direction in response to the increase or decrease of hsp90 induction, depending on the temperature. Suppression of degradation following re-exposure of the cells to 44 degrees C correlated better with induction of hsp90 than hsp70. We infer that sequestration of hsp70 by heat-unfolded receptor is the primary factor opposing degradation, while induction of hsp90 acts to further suppress degradation by accelerating HRC assembly.     

The effect of terpenoid extracts from 15 pine species on the feeding behavioural sequence of the late instars of the pine processionary caterpillar Thaumetopoea pityocampa

The feeding behaviour of the pine processionary (PPC) caterpillar Thaumetopoea pityocampa Den. and Schiff. (Lepidoptera, Thaumetopoeidae) in L3-L4 stages was explored by means of laboratory arena feeding trials and natural substrates. In the bioassays, volatile extracts of 15 pine species, 8 of which are naturally growing in Greece, were incorporated. An analytical model was developed based on the principle of multinomial logit regression with five outcomes on the basis of the behavioural feeding sequence of the caterpillars. The outcomes were the five steps in which the feeding behavioural sequence was decomposed. The model's suitability (MacFadden's rho(2)=0.229, P<10(-4)) was examined when including 10 terpenes that were judged significant through a stepwise canonical discriminant analysis. The proposed model was superior to a random one and the two models resulting from the addition and subtraction of 4 terpenes to the already 10 existing compounds. The most informative model was built on the terpenes caryophyllene oxide, terpinolene, myrcene, germacrene D, eudesmol, limonene, beta-pinene, beta-caryophyllene, alpha-pinene and manoyl oxide. The background terpenes were present in the model and of particular importance. No special behavioural role, either as promoter or inhibitor could be assigned to the individual volatile metabolites, since no constant pattern among behavioural steps was observed. For instance, beta-caryophyllene while acts as promoter of attraction and trial bite it is a suppressor of partial feeding and strongly inhibits complete needle consumption. The monoterpene limonene, on the other hand, seems to be a suppressor of partial and complete feeding. The overall methodological scheme and the analytical modelling tool could be proved a suitable research protocol in unfolding the ecological role of a complex mixture of secondary metabolites. Those who develop safe practical systems can use this.     

Mutations of the coat protein gene of bacteriophage λ that overcome the necessity for the FI gene; the EFi domain

The functions of most of the 10 genes involved in phage λ capsid morphogenesis are well understood. The function of the FI gene is one of the exceptions. Mutants in FI fail to mature and package DNA. The gene product (gpFI) seems to act as a catalyst for the formation of an intermediate in capsid assembly called complex II, which contains a procapsid (an empty capsid precursor), terminase (the enzyme that cleaves the DNA precursor and packages it into the procapsid) and DNA. The mechanism for this stimulation remains unknown. It has also been reported that gpFI appeared to stimulate terminase-mediated cos cleavage, in the absence of procapsids, by increasing enzyme turnover. In comparison with other head-gene mutants, FI mutants are leaky, producing approx. 0.1 phage per infected cell. Some second-site revertants of FI ^? phages, called ‘fin’, that bypass the necessity for gpFI, have been isolated and found to harbour a mutation in the genes that code for the two subunits of terminase. In the course of mapping additional fin mutants, it was discovered that some mapped outside the terminase genes. To localize the mutations, restriction fragments of fin mutant DNAs were subcloned into plasmids and their ability to contribute to fin function was determined by marker-rescue analysis. The location of the fin mutation was further delineated by deletion analysis of a plasmid that was positive for fin. This showed that some fin mutations mapped to a region comprising genes E, D and a portion of C. The sequencing of this entire region in several fin isolates showed that the fin mutations are clustered in a small region of gene E corresponding to a portion of 26 amino acid residues of the coat protein (gpE). We have called this region of the protein the EFi domain. All the mutations result in an increase in positive charge relative to the wild-type protein. These results suggest that DNA maturation and packaging are in part controlled by an interaction between gpFI and capsid gpE.     

Lactobacillus casei Exerts Anti-Proliferative Effects Accompanied by Apoptotic Cell Death and Up-Regulation of TRAIL in Colon Carcinoma Cells

Probiotic microorganisms such as lactic acid bacteria (LAB) exert a number of strain-specific health-promoting activities attributed to their immunomodulatory, anti-inflammatory and anti-carcinogenic properties. Despite recent attention, our understanding of the biological processes involved in the beneficial effects of LAB strains is still limited. To this end, the present study investigated the growth-inhibitory effects of Lactobacillus casei ATCC 393 against experimental colon cancer. Administration of live Lactobacillus casei (as well as bacterial components thereof) on murine (CT26) and human (HT29) colon carcinoma cell lines raised a significant concentration- and time-dependent anti-proliferative effect, determined by cell viability assays. Specifically, a dramatic decrease in viability of colon cancer cells co-incubated with 10⁹ CFU/mL L. casei for 24 hours was detected (78% for HT29 and 52% for CT26 cells). In addition, live L. casei induced apoptotic cell death in both cell lines as revealed by annexin V and propidium iodide staining. The significance of the in vitro anti-proliferative effects was further confirmed in an experimental tumor model. Oral daily administration of 10⁹ CFU live L. casei for 13 days significantly inhibited in vivo growth of colon carcinoma cells, resulting in approximately 80% reduction in tumor volume of treated mice. Tumor growth inhibition was accompanied by L. casei-driven up-regulation of the TNF-related apoptosis-inducing ligand TRAIL and down-regulation of Survivin. Taken together, these findings provide evidence for beneficial tumor-inhibitory, anti-proliferative and pro-apoptotic effects driven by this probiotic LAB strain.     

Copper(II) complexes of diclofenac: Spectroscopic studies and DNA strand breakage

Copper(II) complexes of diclofenac with interesting anti-inflammatory profiles have been prepared and studied by infrared and electronic spectroscopy. In the solid state and in polar and coordinating solvents, all the complexes are solvated binuclear carboxylato-bridged complexes, [Cu(L)²(S)]², where L is monodeprotonated diclofenac and S is the axially bonded solvent. The effect of the copper(II) complexes on the in vitro DNA strand breakeage was studied by agarose gel electrophoresis. Relaxation or double stranded scissions of pDNA were observed leading to the formation of linear pDNA. Treatment of pDNA with high concentrations of these compounds caused a disappearance of pDNA. For the parent drug, sodium diclofenac, no effect on the pDNA was observed. This study presents some indications that the binuclear copper(II) complexes, [Cu(L)²(S)]², could have some relevance in the treatment of tumor cell lines.     

Functional principles of steerable multi‐element probes in insects

Hemipterans, mosquitoes, and parasitic wasps probe in a variety of substrates to find hosts for their larvae or food sources. Probes capable of sensing and precise steering enable insects to navigate through solid substrates without visual information and to reach targets that are hidden deep inside the substrate. The probes belong to non‐related taxa and originate from abdominal structures (wasps) or mouthparts (hemipterans and mosquitoes), but nevertheless share several morphological characteristics. Although the transport function clearly differs (egg laying and acquisition of liquid food), the functional demands on the mechanical behaviour of the probe within the substrate tend to be similar. The probe needs to be thin to limit substrate deformation, and long, in order to attain substantial path lengths or depths. We linked the morphology across taxa to the different functional requirements, to provide insights into the biology of probing insects and the evolution of their probes. Current knowledge of insect probes is spread over many taxa, which offers the possibility to derive general characteristics of insect probing. Buckling during initial puncturing is limited by external support mechanisms. The probe itself consist of multiple (3–6) parts capable of sliding along one another. This multi‐part construction presumably enables advancement and precise three‐dimensional steering of the probe through the substrate with very low net external pushing forces, preventing buckling during substrate penetration. From a mechanical viewpoint, a minimum of three elements is required for 3D steering and volumetric exploration, as realised in the ovipositors of wasps. More elements, such as in six‐element probes of mosquitoes, may enhance friction in soft substrates. Alternatively, additional elements can have functions other than ‘drilling’, such as saliva injection in mosquitoes. Despite the gross similarities, probes show differences in their cross sections, tip morphologies, relative lengths of their elements, and the shape of their interconnections. The hypothesis is that the probe morphology is influenced by the substrate properties, which are mostly unknown. Correlating the observed diversity to substrate‐specific functional demands is therefore currently impossible. We conclude that a multipart probe with sliding elements is highly effective for volumetric substrate probing. Shared functional demands have led to an evolutionary convergence of slender multi‐element probes in disparate insect taxa. To fully understand 3D probing, it is necessary to study the sensory and material properties, as well as the detailed kinematics and dynamics of the various probes in relation to the nature of the selective pressure originating from the species‐specific substrates. Such knowledge will deepen our understanding of probing mechanisms and may support the development of slender, bio‐inspired probes.