The cell shape proteins MreB and MreC control cell morphogenesis by positioning cell wall synthetic complexes

MreB, the bacterial actin homologue, is thought to function in spatially co-ordinating cell morphogenesis in conjunction with MreC, a protein that wraps around the outside of the cell within the periplasmic space. In Caulobacter crescentus, MreC physically associates with penicillin-binding proteins (PBPs) which catalyse the insertion of intracellularly synthesized precursors into the peptidoglycan cell wall. Here we show that MreC is required for the spatial organization of components of the peptidoglycan-synthesizing holoenzyme in the periplasm and MreB directs the localization of a peptidoglycan precursor synthesis protein in the cytosol. Additionally, fluorescent vancomycin (Van-FL) labelling revealed that the bacterial cytoskeletal proteins MreB and FtsZ, as well as MreC and RodA, were required for peptidoglycan synthetic activity. MreB and FtsZ were found to be required for morphogenesis of the polar stalk. FtsZ was required for a cell cycle-regulated burst of peptidoglycan synthesis early in the cell cycle resulting in the synthesis of cross-band structures, whereas MreB was required for lengthening of the stalk. Thus, the bacterial cytoskeleton and cell shape-determining proteins such as MreC, function in concert to orchestrate the localization of cell wall synthetic complexes resulting in spatially co-ordinated and efficient peptidoglycan synthetic activity.     

Structure of Spa15, a type III secretion chaperone from Shigella flexneri with broad specificity

Type III secretion (TTS) systems are used by many Gram-negative pathogens to inject virulence proteins into the cells of their hosts. Several of these virulence effectors require TTS chaperones that maintain them in a secretion-competent state. Whereas most chaperones bind only one effector, Spa15 from the human pathogen Shigella flexneri and homologous chaperones bind several seemingly unrelated effectors, and were proposed to form a special subgroup. Its 1.8 A crystal structure confirms this specific classification, showing that Spa15 has the same fold as other TTS effector chaperones, but forms a different dimer. The presence of hydrophobic sites on the Spa15 surface suggests that the different Spa15 effectors all possess similar structural elements that can bind these sites. Furthermore, the Spa15 structure reveals larger structural differences between class I chaperones than previously anticipated, which does not support the hypothesis that chaperone-effector complexes are structurally conserved and function as three-dimensional secretion signals.     

Studies on the factors influencing larval settlement in Balanus amphitrite and Mytilus galloprovincialis

Understanding of factors influencing settlement(attachment and metamorphosis) of marine invertebratelarvae is of great importance in aquaculture andcontrol of biofouling. The influence of two factors onsettlement of larvae was assessed from two separateinvestigations: 1, the influence of age (endogenousfactor) on cyprids of the barnacle Balanusamphitrite; and 2, the influence of a microbial film(exogenous factor) on pediveligers of the mussel Mytilus galloprovincialis.The settlement response of cypris larvae of B.amphitrite was found to be age-dependent. Oldercyprids responded more readily to settlement factorsthan newly molted ones. In M.galloprovincialis, competent pediveligers settled inresponse to a microbial filmed surface but not toan unfilmed surface. Moreover, a factor with MW of lessthan 5000 dalton, derived from culture medium of abacterial strain C1.1 (Pseudomonas-Alteromonasgroup), induced the settlement of M. galloprovincialis larvae.Thus, marine invertebrate larvae may require a periodof competence acquisition, during which they arepoorly responsive to settlement inducers. Uponacquisition of competence, larvae readily respondto external cues (e.g. microbial film, bacterialextracellular products).     

Computational study of radiation chemical processing in comet nuclei

Cometary nuclei have been exposed to high levels of ionizing radiation since their formation. We present here some results of a computer model calculation of the effect of ionizing radiation on cometary material. The external (cosmic rays) and internal (embedded radionuclides) contributions in the processing of cometary nuclei are considered. As a first approximation we have used the available kinetic data of the liquid water system to model the radiation effects in a frozen cometary environment. Out data suggest that massive radiation chemical processing due to cosmic rays may have taken place only in the outer layers of comets. The internal contribution of radionuclides to the radiation processing of comet cores seems to be modest. Therefore, comets could be carriers of intact homochiral biomolecules.Part of this work was carried out during a leave at the Laboratory of Chemical Evolution.     

Incorporation of ethidium bromide in the Drosophila salivary gland approached by microspectrofluorometry: evidence for the presence of both free and bound dye in the nuclei of cells in viable conditions

The incorporation of 10^–6 M ethidium bromide (EB) was studied in viable Drosophila melanogaster salivary glands with a spatial resolution reaching a few µm³, using a confocal laser microspectrofluorometer designed for spectral analysis. Spectra were recorded with the 514 nm Argon laser line during excitation times of 1 second (20 µW on the preparation) at 5 min intervals for 30 or 60 min, either at points in determined cell sites or serially throughout the cells. The fluorescence intensity time-course indicated that the EB intake was not an all-or-none process, but rather a graded, sensitive indicator of the functional state of the cell. On the micrometer scale, the cytoplasm behaved as an homogeneous substrate with the fluorescence intensity depending on EB intake and intracellular diffusion. In the nucleus, however, localized enhancement of the emission intensity was observed. Spectral analysis allowed us to characterize the interactions. The mean values of λ _max in the cytoplasm (600 nm), in the nucleus (601 nm) and outside the glands (602 nm) were less than for free EB in aqueous solution (630 nm); values of full width at half maximum were between 92 and 96 nm, which is much lower than the 120 nm observed for free EB. The recorded spectra were analyzed using a linear combination of two spectral models, namely free and DNA intercalated EB. In the nucleus, the free EB model spectra was found to represent up to 10% of the recorded spectra whereas it was near zero in the cytoplasm. The present data suggest that the intranuclear concentration of free EB (allowing for its lower fluorescence quantum yield) might be at least equal to that of the bound EB.     

Interface processes between protected and unprotected areas: A global review and ways forward

Land‐use changes and the expansion of protected areas (PAs) have amplified the interaction between protected and unprotected areas worldwide. In this context, ‘interface processes' (human–nature and cross‐boundary interactions inside and around PAs) have become central to issues around the conservation of biodiversity and ecosystem services. This scientific literature review aimed to explore current knowledge and research gaps on interface processes regarding terrestrial PAs. At first, 3,515 references related to the topic were extracted through a standardized search on the Web of Science and analyzed with scientometric techniques. Next, a full‐text analysis was conducted on a sample of 240 research papers. A keyword analysis revealed a wide diversity of research topics, from ‘pure' ecology to sociopolitical research. We found a bias in the geographical distribution of research, with half the papers focusing on eight countries. Additionally, we found that the spatial extent of cross‐boundary interactions was rarely assessed, preventing any clear delimitation of PA interactive zones. In the 240 research papers we scanned, we identified 403 processes that were studied. The ecological effects of PAs were well documented and appeared to be positive overall. In contrast, the effects of PAs on local communities were understudied and, according to the literature focusing on these, were very variable according to local contexts. Our findings highlight key research advances on interface processes, especially regarding the ecological outcomes of PAs, the influence of human activities on biodiversity, and PA governance issues. In contrast, main knowledge gaps concern the spatial extent of interactive zones, as well as the interactions between local people and conservation actions and how to promote synergies between them. While the review was limited to terrestrial PAs, its findings allow us to propose research priorities for tackling environmental and socioeconomic challenges in the face of a rapidly changing world.     

Arabidopsis RECEPTOR-LIKE PROTEIN30 and Receptor-Like Kinase SUPPRESSOR OF BIR1-1/EVERSHED Mediate Innate Immunity to Necrotrophic Fungi

Effective plant defense strategies rely in part on the perception of non-self determinants, so-called microbe-associated molecular patterns (MAMPs), by transmembrane pattern recognition receptors leading to MAMP-triggered immunity. Plant resistance against necrotrophic pathogens with a broad host range is complex and yet not well understood. Particularly, it is unclear if resistance to necrotrophs involves pattern recognition receptors. Here, we partially purified a novel proteinaceous elicitor called SCLEROTINIA CULTURE FILTRATE ELICITOR1 (SCFE1) from the necrotrophic fungal pathogen Sclerotinia sclerotiorum that induces typical MAMP-triggered immune responses in Arabidopsis thaliana. Analysis of natural genetic variation revealed five Arabidopsis accessions (Mt-0, Lov-1, Lov-5, Br-0, and Sq-1) that are fully insensitive to the SCFE1-containing fraction. We used a forward genetics approach and mapped the locus determining SCFE1 sensitivity to RECEPTOR-LIKE PROTEIN30 (RLP30). We also show that SCFE1-triggered immune responses engage a signaling pathway dependent on the regulatory receptor-like kinases BRASSINOSTEROID INSENSITIVE1-ASSOCIATED RECEPTOR KINASE1 (BAK1) and SUPPRESSOR OF BIR1-1/EVERSHED (SOBIR1/EVR). Mutants of RLP30, BAK1, and SOBIR1 are more susceptible to S. sclerotiorum and the related fungus Botrytis cinerea. The presence of an elicitor in S. sclerotiorum evoking MAMP-triggered immune responses and sensed by RLP30/SOBIR1/BAK1 demonstrates the relevance of MAMP-triggered immunity in resistance to necrotrophic fungi.     

Inhibition of DNA damage repair by artificial activation of PARP with siDNA

One of the major early steps of repair is the recruitment of repair proteins at the damage site, and this is coordinated by a cascade of modifications controlled by phosphatidylinositol 3-kinase-related kinases and/or poly (ADP-ribose) polymerase (PARP). We used short interfering DNA molecules mimicking double-strand breaks (called Dbait) or single-strand breaks (called Pbait) to promote DNA-dependent protein kinase (DNA-PK) and PARP activation. Dbait bound and induced both PARP and DNA-PK activities, whereas Pbait acts only on PARP. Therefore, comparative study of the two molecules allows analysis of the respective roles of the two signaling pathways: both recruit proteins involved in single-strand break repair (PARP, XRCC1 and PCNA) and prevent their recruitment at chromosomal damage. Dbait, but not Pbait, also inhibits recruitment of proteins involved in double-strand break repair (53BP1, NBS1, RAD51 and DNA-PK). By these ways, Pbait and Dbait disorganize DNA repair, thereby sensitizing cells to various treatments. Single-strand breaks repair inhibition depends on direct trapping of the main proteins on both molecules. Double-strand breaks repair inhibition may be indirect, resulting from the phosphorylation of double-strand breaks repair proteins and chromatin targets by activated DNA-PK. The DNA repair inhibition by both molecules is confirmed by their synthetic lethality with BRCA mutations.