Combining μX-ray fluorescence, μXANES and μXRD to shed light on Zn2+ cations in cartilage and meniscus calcifications

We aimed to examine the presence of Zn, a trace element, in osteoarthritis (OA) cartilage and meniscus from patients undergoing total knee joint replacement for primary OA. We mapped Ca²⁺ and Zn²⁺ at the mesoscopic scale by X-ray fluorescence microanalysis (μX-ray) to determine the spatial distribution of the 2 elements in cartilage, μX-ray absorption near edge structure spectroscopy to identify the Zn species, and μX-ray diffraction to determine the chemical nature of the calcification. Fourier transform infrared spectroscopy was used to determine the chemical composition of cartilage and meniscus. Ca²⁺ showed a heterogeneous spatial distribution corresponding to the calcifications within cartilage (or meniscus) or at their surface. At least 2 Zn²⁺ species were present: the first may correspond to Zn embedded in protein (different Zn metalloproteins are known to prevent calcification in biological tissues), and the second may be associated with a Zn trap in or at the surface of the calcification. Calcification present in OA cartilage may significantly modify the spatial distribution of Zn; part of the Zn may be trapped in the calcification and may alter the associated biological function of Zn metalloproteins.     

Embryonic exposure to predator odour modulates visual lateralization in cuttlefish

Predation pressure acts on the behaviour and morphology of prey species. In fish, the degree of lateralization varies between high- and low-predation populations. While lateralization appears to be widespread in invertebrates, we do not know whether heredity and early experience interact during development as in vertebrates. Here we show, for the first time, that an exposure to predator odour prior to hatching modulates visual lateralization in newly hatched cuttlefish. Only cuttlefish that have been exposed to predator odour display a left-turning bias when tested with blank seawater in a T-shaped apparatus. Exposure to predator odour all the incubation long could appear as an acute predictor of a high-predation surrounding environment. In addition, cuttlefish of all groups display a left-turning preference when tested with predator odour in the apparatus. This suggests the ability of cuttlefish to innately recognize predator odour. To our knowledge, this is the first clear demonstration that lateralization is vulnerable to ecological challenges encountered during embryonic life, and that environmental stimulation of the embryo through the olfactory system could influence the development of subsequent visual lateralization.     

Imitating the cost of males: A hypothesis for coexistence of all‐female sperm‐dependent species and their sexual host

All‐female sperm‐dependent species are particular asexual organisms that must coexist with a closely related sexual host for reproduction. However, demographic advantages of asexual over sexual species that have to produce male individuals could lead both to extinction. The unresolved question of their coexistence still challenges and fascinates evolutionary biologists. As an alternative hypothesis, we propose those asexual organisms are afflicted by a demographic cost analogous to the production of males to prevent exclusion of the host. Previously proposed hypotheses stated that asexual individuals relied on a lower fecundity than sexual females to cope with demographic advantage. In contrast, we propose that both sexual and asexual species display the same number of offspring, but half of asexual individuals imitate the cost of sex by occupying ecological niches but producing no offspring. Simulations of population growth in closed systems under different demographic scenarios revealed that only the presence of nonreproductive individuals in asexual females can result in long‐term coexistence. This hypothesis is supported by the fact that half of the females in some sperm‐dependent organisms did not reproduce clonally.     

Salmo macrostigma (Teleostei,Salmonidae): Nothing more than a brown trout (S. trutta) lineage?

We examined specimens of the macrostigma trout Salmo macrostigma, which refers to big black spots on the flanks, to assess whether it is an example of taxonomic inflation within the brown trout Salmo trutta complex. Using new specimens, publicly available data and a mitogenomic protocol to amplify the control and cytochrome b regions of the mitochondrial genome from degraded museum samples, including one syntype specimen, the present study shows that the macrostigma trout is not a valid species. Our results suggest the occurrence of a distinct evolutionary lineage of S. trutta in North Africa and Sicily. The name of the North African lineage is proposed for this lineage, which was found to be sister to the Atlantic lineage of brown trout, S. trutta.     

Two subclasses of guanine exchange factor (GEF) domains revealed by comparison of activities of chimeric genes constructed from CDC25, SDC25 and BUD5 in Saccharomyces cerevisiae

Guanine Exchange Factor (GEF) activity for Ras proteins has been associated with a conserved domain in Cdc25p, Sdc25p in Saccharomyces cerevisiae and several other proteins recently found in other eukaryotes. We have assessed the structure-function relationships between three different members of this family in S. cerevisiae, Cdc25p, Sdc25p and Bud5p. Cdc25p controls the Ras pathway, whereas Bud5p controls bud site localization. We demonstrate that the GEF domain of Sdc25p is closely related to that of Cdc25p. We first constructed a thermosensitive allele of SDC25 by specifically altering amino acid positions known to be changed in the cdc25-1 mutation. Secondly, we constructed three chimeric genes from CDC25 and SDC25, the products of which are as active in the Ras pathway as are the wild-type proteins. In contrast, similar chimeras made between CDC25 and BUD5 lead to proteins that are inactive both in the Ras and budding control pathways. This difference in the ability of chimeric proteins to retain activity allows us to define two subclasses of structurally different GEFs: Cdc25p and Sdc25p are Ras-specific GEFs, and Bud5p is a putative GEF for the Rsr1/Bud1 Rap-like protein.     

Identification of potent RORβ modulators: Scaffold variation

We sought to develop RORβ-selective probe molecules in order to investigate the function of the receptor in vitro and in vivo and its role in the pathophysiology of disease. To accomplish this, we modified a potent dual RORβ/RORγ inverse agonist from the primary literature with the goal of improving selectivity for RORβ vs RORγ. Truncation of the Western portion of the molecule ablated activity at RORγ and led to a potent series of RORβ modulators. Continued exploration of this series investigated alternate replacement cores for the aminothiazole ring. Numerous suitable replacements were found during the course of our SAR investigations and are reported herein.     

Antibacterial activities of mono-, di- and tri-substituted triphenylamine-based phosphonium ionic liquids

We report the synthesis of new mono, di and tri phosphonium ionic liquids and the evaluation of their antibacterial activities on both Gram-positive and Gram-negative bacteria from the ESKAPE-group. Among the molecules synthesized some of them reveal a strong bactericidal activity (MIC?=?0.5?mg/L) for Gram-positive bacteria (including resistant strains) comparable to that of standard antibiotics. A comparative Gram positive and Gram negative antibacterial activities shows that the nature of counter-ion has no significant effects. Interestingly, the increase of phosphonium lateral chains (from 4 to 8 carbons) results in a decrease of antibacterial activities. However, the increase of the spacer length has a positive influence on the activity on both Gram-positive and Gram-negative bacteria except for E. aerogenes. Finally, the increased charge density has no effect on the Gram-positive antibacterial activities (MIC between 2 and 4?mg/L) but seems to attenuate (except for P. aeruginosa) the discrimination between Gram-positive and Gram-negative. Overall these results suggest a unique mechanism of action of these triphenylamine-phosphonium ionic liquid derivatives.     

Syntheses and antiproliferative activities of rebeccamycin analogues bearing two 7-azaindole moieties

As a part of structure-activity relationship studies on rebeccamycin analogues, compounds containing two aza-indole moieties were synthesized bearing either a methyl group or a hydrogen atom on the imide nitrogen. The azaindole substructures were expected to enhance the cytotoxicity toward tumor cell lines through stronger hydrogen bonding with the target enzyme(s). The cytotoxicities of compounds 8, 10 and 19 against a panel of tumor cell lines were examined and compared with those of rebeccamycin, dechlorinated rebeccamycin 2 and N-methylated analogue A. Their effect on the L1210 cell cycle was also evaluated. Compound 19, having an imide NH function had the strongest cytotoxicity towards L1210 cells and induced the largest accumulation of cells in the G2+M phases of the cell cycle. In contrast to their non-aza analogues, which were cytotoxic for all the cell lines tested, diaza compounds 10 and 19 showed selectivity for some cell lines.     

Chloroplast Activity and 3′phosphadenosine 5′phosphate Signaling Regulate Programmed Cell Death in Arabidopsis

Programmed cell death (PCD) is a crucial process both for plant development and responses to biotic and abiotic stress. There is accumulating evidence that chloroplasts may play a central role during plant PCD as for mitochondria in animal cells, but it is still unclear whether they participate in PCD onset, execution, or both. To tackle this question, we have analyzed the contribution of chloroplast function to the cell death phenotype of the myoinositol phosphate synthase1 (mips1) mutant that forms spontaneous lesions in a light-dependent manner. We show that photosynthetically active chloroplasts are required for PCD to occur in mips1, but this process is independent of the redox state of the chloroplast. Systematic genetic analyses with retrograde signaling mutants reveal that 3′-phosphoadenosine 5′-phosphate, a chloroplast retrograde signal that modulates nuclear gene expression in response to stress, can inhibit cell death and compromises plant innate immunity via inhibition of the RNA-processing 5′-3′ exoribonucleases. Our results provide evidence for the role of chloroplast-derived signal and RNA metabolism in the control of cell death and biotic stress response.Programmed cell death (PCD) is a universal process in multicellular organisms, contributing to the controlled and active degradation of the cell. In plants, PCD is required for processes as diverse as development, self-incompatibility, and stress response. One well-documented example is the induction of PCD upon pathogen attack, allowing the confinement of the infection, and resistance of the plant. The signaling events leading to the onset of PCD have been extensively studied: pathogen recognition triggers activation of mitogen-activated protein kinase cascades, as well as production of reactive oxygen species (ROS) and salicylic acid (SA), which lead to a hypersensitive response (Coll et al., 2011).From a cellular point of view, several classes of plant PCD have been described and compared with the ones found in animal cells (van Doorn, 2011). PCD is thought to have evolved independently in plants and animals, and genes underlying these mechanisms are therefore poorly conserved between the two kingdoms. However, most cellular features are conserved between plant and animal PCD that are both characterized by cell shrinkage, chromatin condensation, DNA laddering, mitochondria permeabilization, and depolarization (Dickman and Fluhr, 2013). In animal cells, mitochondria play a central role in the regulation of apoptosis (Czabotar et al., 2014; Mariño et al., 2014), and this role is likely shared between the two kingdoms (Lord and Gunawardena, 2012). That said, additional mitochondria-independent PCD pathways have clearly evolved in plants.Genetic approaches have greatly contributed to our understanding of cellular pathways governing PCD in plants. For example, the isolation of lesion mimic mutants (LMMs), in which cell death occurs spontaneously, has allowed the identification of several negative regulators of cell death (for review, see Bruggeman et al., 2015b). Interestingly, lesion formation is light dependent in several of these mutants, which include one of the best characterized LMMs—lesions simulating disease1 (lsd1; Dietrich et al., 1994). The LSD1 protein is required for plant acclimation to excess excitation energy (Mateo et al., 2004): when plants are exposed to excessive amounts of light, the redox status of the plastoquinone pool in the chloroplastic electron transfer chain is thought to influence LSD1-dependent signaling to modulate cell death (Mühlenbock et al., 2008). Additionally, we have previously identified the myoinositol phosphate synthase1 (mips1) mutant as a LMM, in which lesion formation is also light dependent (Meng et al., 2009). This mutant is deficient in the myoinositol (MI) phosphate synthase that catalyzes the first committed step of MI biosynthesis and displays pleiotropic defects such as reduced root growth, abnormal vein development, and spontaneous cell death on leaves, together with severe growth reduction after lesions begin to develop (Meng et al., 2009; Donahue et al., 2010). The light-dependent PCD in the mips1 mutant, as observed for lsd1, suggests that chloroplasts may play a role in the MI-dependent cell death regulation. Accumulating evidence suggests that chloroplasts may play a central role in PCD regulation like mitochondria in animal cells (Wang and Bayles, 2013). First, as described in the case of lsd1, excess light energy received by the chloroplast can function as a trigger for PCD. Furthermore, singlet oxygen (¹O₂), a ROS, can activate the EXECUTER1 (EX1) and EX2 proteins in the chloroplasts to initiate PCD (Lee et al., 2007). Likewise, ROS generated by chloroplasts play a major role for PCD onset during nonhost interaction between tobacco (Nicotiana tabacum) and Xanthomonas campestris (Zurbriggen et al., 2009). Finally, functional chloroplasts have also been shown to be required for PCD in cell suspensions (Gutierrez et al., 2014) and in a number of LMMs (Mateo et al., 2004; Meng et al., 2009; Bruggeman et al., 2015b). Thus, chloroplasts are now recognized as important components of plant defense response against pathogens (Stael et al., 2015) and are proposed to function with mitochondria in the execution of PCD (Van Aken and Van Breusegem, 2015). However, the exact signaling and metabolic contribution of chloroplasts to PCD remain to be elucidated. Furthermore, cross talk between chloroplasts and mitochondria does occur, such as during photorespiration (Sunil et al., 2013), but whether such communication functions sequentially or in parallel in the control of PCD remains to be determined (Van Aken and Van Breusegem, 2015).To further investigate how chloroplasts contribute to the regulation of cell death, we performed both forward and reverse genetics on the mips1 mutant. An extragenic secondary mutation in divinyl protochlorophyllide 8-vinyl reductase involved in chlorophyll biosynthesis leads to chlorophyll deficiency that abolishes the mips1 cell death phenotype, as do changes in CO₂ availability. These findings provide evidence for a link between photosynthetic activity and PCD induction in mips1. Additionally, we investigated the contribution of several retrograde signaling pathways (Chan et al., 2015) to the control of PCD in mips1. This process was independent of GENOMES UNCOUPLED (GUN) and EX signaling pathways, but we found that the SAL1-PAP_XRN retrograde signaling pathway inhibits cell death as well as basal defense reactions in Arabidopsis (Arabidopsis thaliana).