Amino acid incorporation by a natural population of Oscillatoria rubescens. A microautoradiographic study

Abstract The incorporation of a very low concentration (0.015 μM) of an [³H]amino acid mixture was measured for a natural population of Oscillatoria rubescens DC in samples from a eutrophic lake, Lake Nantua (France).
The kinetics of amino acid incorporation in the size fraction ⩾12 μ m showed that uptake was fast and that the maximum was reached after 4 h.
Microautoradiography demonstrated that Oscillatoria rubescens is able to utilize for protein synthesis an external pool of amino acids whose [³H] label becomes distributed generally throughout the cell.     

Population responses of roe deer to the recolonization of the French Vercors by wolves

In a context of changing carnivore populations worldwide, it is crucial to understand the consequences of these changes for prey populations. The recolonization by wolves of the French Vercors mountain range and the long-term monitoring (2001–2017) of roe deer in this area provided a unique opportunity to assess the effects of wolves on this prey. Roe deer was the main prey of wolves in the west Vercors mountain range during this recolonization. We compared roe deer abundance and fawn body mass in two contrasted areas of a wolf pack territory: a central area (core of the territory characterized by an intense use by wolves) and a peripheral area (used more occasionally). Roe deer population growth rates were lower in the central area between 2001 and 2006, resulting in a decline in roe deer abundance. Roe deer abundance substantially dropped in the two study areas after an extremely severe winter but the abundance of roe deer in the central area facing with wolves was slower to recover and remained at lower abundance levels for 6 years. Fawn body mass was consistently lower in the central area, varied similarly as roe deer abundance, and was not influenced by weather conditions or red deer population abundance. Altogether, the effects of wolves on roe deer in the central area occurred during a 10-year period following the establishment of wolves, through the interplay between wolf predation (before wolves started preying on red deer), harsh winter conditions and possibly naivety of prey to this recolonizing predator.     

Mercury and non-protein thiol compounds in the seagrass Posidonia oceanica

Mercury concentrations, non-protein thiol levels and the enzyme activities of glutathione-S-transferase (GST) were measured in the blades and sheaths of the marine phanerogam Posidonia oceanica. The seagrass was collected in January and June and at three sites: the Bay of Rosignano (Italy) known for its mercury contamination, the north of the Lérins islands (Bay of Cannes, France), the Bay of Tonnara (Corsica, France). The two latter sites are considered as free of any known industrial inputs. Mercury concentrations and GST activities in both tissues were always higher in samples from Rosignano, particularly in June. Non-protein thiol levels were significantly higher in the blades than in the sheaths of P. oceanica from Tonnara and Lérins. In contrast, at Rosignano, the sheaths presented a significantly higher non-protein thiol concentration than the blades, particularly in June. Levels in the sheaths appeared to increase with the degree of pollution. Western Blot performed on sheaths of P. oceanica collected in June at Rosignano and Lérins revealed a characteristic band of GSTs at 31 kDa, proving the presence of the GST enzyme in this tissue. Mercury seemed to exert an influence upon non-protein thiol metabolism, including GST induction, in P. oceanica collected from the NW Mediterranean.     

Genetic Structure and Internal Rearrangements of Stable Merodiploids from BACILLUS SUBTILIS Strains Carrying the trpE26 Mutation

Transformation and transduction to tryptophan independence of strains of Bacillus subtilis carrying the "trpE26" chromosomal aberrations (a translocation and an inversion) with a "normal" 168 type strain as donor induce a tandem duplication of the thrA-ilvA region of the chromosome. The clones possessing this unstable duplication segregate besides the Trp^- some stable Trp⁺ cells which retain only part of the duplication (the trpE-ilvA region) in nontandem configuration. Such clones may also be produced directly during the crosses. The genetic map of these clones (designated as class I stable merodiploids) was constructed: they possess the tranlocation and the inversion of the trpE26 parental strain. Another type of stable Trp⁺ clones (class II) also appears, although more rarely, in similar crosses. Studies on their genetic structure revealed that they are haploid for the trpE-ilvA region and carry a nontandem duplication of the thrA-trpE region. In these clones the cysB-tre region has the orientation of the 168 type strain. The duplications in both classes are stable, that of class I being more stable than that of class II where loss of one copy of the thrA-trpE region leads to about 1% haploid cells. Detailed genetic studies on heterozygous clones from both classes have shown exchange of alleles between copies of the nontandem duplications. Models are proposed for the formation of each class of merodiploids and for recombination events taking place in them. These models imply recombination at sequences of intrachromosomal homology and (or) introduction of heterologous juncions ("novel joints") by transformation or transduction.     

Conventional Pap smear and liquid-based cytology as screening tools in low-resource settings in Latin America: experience of the Latin American screening study

OBJECTIVE: To evaluate the performance of the conventional Pap test and liquid-based cytology (LBC) in an ongoing multicenter trial testing optional screening tools (cytology, screening colposcopy, visual inspection with acetic acid, visual inspection with Lugol's Iodine, cervicography and Hybrid Capture II [HCII] (Digene Brazil, S?o Paulo, Brazil) conventional and self-sampling), for cervical cancer in Brazil and Argentina. STUDY DESIGN: A cohort of 12,107 women attending four clinics (Campinas, S?o Paulo, Porto Alegre, Buenos Aires) were randomized into the 8 diagnostic arms. Women testing positive with any of the tests were referred for colposcopy, and cervical biopsies were used as the gold standard to assess performance characteristics of the diagnostic tests. Conventional Pap smears were sampled by all clinics (n = 10,240), and LBC (Autocyte PREP, [TriPath Imaging, Burlington, North Carolina, U.S.A.], n=320, and DNA-Citoliq [Digene Brazil], n =1,346) was performed by 1 of the clinics. RESULTS: Conventional Pap smears showed no squamous intraepithelial lesions (normal) in 8,946 (87.4%) and LBC in 1,373 (82.4%). Using high grade squamous intraepithelial lesions (HSIL) as the cutoff, Pap smears predicted high grade (cervical intraepithelial neoplasia [CIN] 3) with OR 63.0 (95% CI, 36.90-107.70), standard error (SE) 59%, SP 97.8%, positive predictive value (PPV) 68.1% and negative predictive value (NPV) 96.7%. The same figures for Autocyte PREP were: OR 9.0 (95% CI, 2.43-33.24), sensitivity (SE) 33.3%, specificity (SP) 100%, PPV 100% and negative PV (NPV) 88.8%. DNA-Citoliq detected CIN 3 as follows: OR 11.8 (95% CI 2.60-53.26), SE 40.0%, SP 94.6%, PPV 40.0% and NPV 94.6%. Lowering the cutoff to low grade squamous intraepithelial lesions increased SE and NPV but compromised SP and PPV. The detection rates for high grade lesions after an atypical squamous cells of undetermined significance diagnosis were similar with the 3 techniques. In our settings, the 3 methods of cervical cytology were slightly different in performance. The conventional Pap smear had the highest SE, while Autocyte PREP had 100% SP and PPV in detecting CIN3 with the HSIL cutoff. All 3 tests had lower SE but higher SP as compared to HCII.     

D-myo-inositol-1,4,5-trisphosphate and adenophostin mimics: importance of the spatial orientation of a phosphate group on the biological activity

Three different routes for the synthesis of heterocyclic analogues of the second messenger D-myo-inositol-1,4,5-trisphosphate (InsP(3)) and the natural adenophostins, starting from allyl D-xyloside are described. The two diastereoisomers at C-2 of new compounds, which we named xylophostins, were obtained. The preliminary biological studies shows that the presence of the adenine residue has a beneficial effect on the affinity for the receptor. The low potency of one of the two diastereoisomeric compounds shows that the configuration of the carbon bearing the non-vicinal phosphate group is an important requirement for a high affinity to the receptor. These results provide evidence for the existence of a binding pocket for the adenine ring nearby the InsP(3) binding site. The consequence of these stabilizing interactions should be to place the phosphate group in a suitable position to perfectly mimic InsP(3) in the more active diastereoisomer. Obviously, in the other diastereoisomer, the phosphate cannot accommodate the same orientation, thus explaining the low affinity. The existence of such a binding pocket for adenine is in line with the high potency of adenophostins.     

Compromising the unfolded protein response induces autophagy-mediated cell death in multiple myeloma cells

Objective

To determine whether the Unfolded Protein Response (UPR) sensors (PERK, ATF6 and IRE-1) can be targeted to promote death of Multiple Myeloma (MM) cells.

Methods

We have knocked-down separately each UPR stress sensor in human MM cell lines using RNA interference and followed MM cell death by monitoring the membrane, mitochondrial and nuclear alterations. Involvement of caspases in MM cell death consecutive to UPR sensor knock-down was analyzed by western blotting, measurement of their enzymatic activity using fluorigenic substrates and susceptibility to a pan-caspase inhibitor. Activation of the autophagic process was measured directly by detection of autophagosomes (electronic microscopy), monodansylcadaverine staining, production of the cleaved form of the microtubule-associated protein 1A/1B light chain 3 (LC3) and indirectly by analyzing the impact of pharmacological inhibitors of autophagy such as 3MA and bafilomycin A1.

Results

We show that extinction of a single UPR stress sensor (PERK) induces a non-apoptotic form of cell death in MM cells that requires autophagy for its execution. We also show that this cytotoxic autophagic process represses the apoptosis program by reducing the cytosolic release of the apoptogenic factors Smac/DIABLO and cytochrome c.

Interpretation

Altogether our findings suggest that autophagy can contribute to execution of death in mammalian cells that are exposed to mild ER stress. They also suggest that the autophagic process can regulate the intrinsic apoptotic pathway by inhibiting production of death effectors by the mitochondria, thus preventing formation of a functional apoptosome. Altogether these findings give credit to the idea that UPR sensors can be envisaged as therapeutic targets for the treatment of MM.