Applications of chlorophyll fluorescence can improve crop production strategies: an examination of future possibilities

Chlorophyll fluorescence has been routinely used for many years to monitor the photosynthetic performance of plants non-invasively. The relationships between chlorophyll fluorescence parameters and leaf photosynthetic performance are reviewed in the context of applications of fluorescence measurements to screening programmes which seek to identify improved plant performance. The potential role of chlorophyll fluorescence imaging in increasing both the sensitivity and throughput of plant screening programmes is examined. Finally, consideration is given to possible specific applications of chlorophyll fluorescence for screening of plants for tolerance to environmental stresses and for improvements in glasshouse production and post-harvest handling of crops.     

Electron microscopy visualization of DNA-protein complexes formed by Ku and DNA ligase IV

The repair of DNA double-stranded breaks (DSBs) is essential for cell viability and genome stability. Aberrant repair of DSBs has been linked with cancer predisposition and aging. During the repair of DSBs by non-homologous end joining (NHEJ), DNA ends are brought together, processed and then joined. In eukaryotes, this repair pathway is initiated by the binding of the ring-shaped Ku heterodimer and completed by DNA ligase IV. The DNA ligase IV complex, DNA ligase IV/XRRC4 in humans and Dnl4/Lif1 in yeast, is recruited to DNA ends in vitro and in vivo by an interaction with Ku and, in yeast, Dnl4/Lif1 stabilizes the binding of yKu to in vivo DSBs. Here we have analyzed the interactions of these functionally conserved eukaryotic NHEJ factors with DNA by electron microscopy. As expected, the ring-shaped Ku complex bound stably and specifically to DNA ends at physiological salt concentrations. At a ratio of 1 Ku molecule per DNA end, the majority of DNA ends were occupied by a single Ku complex with no significant formation of linear DNA multimers or circular loops. Both Dnl4/Lif1 and DNA ligase IV/XRCC4 formed complexes with Ku-bound DNA ends, resulting in intra- and intermolecular DNA end bridging, even with non-ligatable DNA ends. Together, these studies, which provide the first visualization of the conserved complex formed by Ku and DNA ligase IV at juxtaposed DNA ends by electron microscopy, suggest that the DNA ligase IV complex mediates end-bridging by engaging two Ku-bound DNA ends.     

TGF-β1 signaling plays a dominant role in the crosstalk between TGF-β1 and the aryl hydrocarbon receptor ligand in prostate epithelial cells

Crosstalk between the aryl hydrocarbon receptor (AhR) and transforming growth factor-β1 (TGF-β1) signaling has been observed in various experimental models. However, both molecular mechanism underlying this crosstalk and tissue-specific context of this interaction are still only partially understood. In a model of human non-tumorigenic prostate epithelial cells BPH-1, derived from the benign prostatic hyperplasia, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) persistently activates the AhR signaling pathway and induces expression of xenobiotic metabolizing enzymes, such as CYP1A1 or CYP1B1. Here we demonstrate that TGF-β1 suppresses the AhR-mediated gene expression through multiple mechanisms, involving inhibition of AhR expression and down-regulation of nuclear AhR, via a SMAD4-dependent pathway. In contrast, TCDD-induced AhR signaling does not affect either TGF-β1-regulated gene expression or epithelial-to-mesenchymal transition. These observations suggest that, in the context of prostate epithelium, TGF-β1 signaling plays a dominant role in the crosstalk with AhR signaling pathway. Given the importance of TGF-β1 signaling in regulation of prostate epithelial tissue homeostasis, as well as the recently revealed role of AhR in prostate development and tumorigenesis, the above findings contribute to our understanding of the mechanisms underlying the crosstalk between the two signaling pathways in the prostate-specific context.     

Synthesis and biological activity of benzo-fused 7-deazaadenosine analogues. 5- and 6-substituted 4-amino- or 4-alkylpyrimido[4,5-b]indole ribonucleosides

Two series of new 4-aminopyrimido[4,5-b]indole ribonucleosides bearing phenyl or hetaryl group at position 5 or 6 have been prepared by Suzuki or Stille cross-coupling reactions employing X-Phos ligand with (het)arylboronic acids or stannanes. A series of 4-substituted nucleosides has been also prepared by Pd-catalyzed cross-couplings or nucleophilic substitution. Some of these compounds displayed moderate antiviral activities against HCV and dengue viruses.     

Exploring Leptin Antagonism in Ophthalmic Cell Models

Background

Emerging evidence suggests that angiogenic and pro-inflammatory cytokine leptin might be implicated in ocular neovascularization. However, the potential of inhibiting leptin function in ophthalmic cells has never been explored. Here we assessed mitogenic, angiogenic, and signaling leptin activities in retinal and corneal endothelial cells and examined the capability of a specific leptin receptor (ObR) antagonist, Allo-aca, to inhibit these functions.

Methods and Results

The experiments were carried out in monkey retinal (RF/6A) and bovine corneal (BCE) endothelial cells. Leptin at 50-250 ng/mL stimulated the growth of both cell lines in a dose-dependent manner. The maximal mitogenic response (35±7 and 27±3% in RF6A and BCE cells, respectively) was noted at 24 h of 250 ng/mL leptin treatments. Leptin-dependent proliferation was reduced to base levels with 10 and 100 nM Allo-aca in BCE and RF6A cells, respectively. In both cell lines, leptin promoted angiogenic responses, with the maximal increase in tube formation (163±10 and 133±8% in RF6A and BCE cultures, respectively) observed under a 250 ng/mL leptin treatment for 3 h. Furthermore, in both cell lines 250 ng/mL leptin modulated the activity or expression of several signaling molecules involved in proliferation, inflammatory activity and angiogenesis, such as STAT3, Akt, and ERK1/2, COX2, and NFκB. In both cell lines, leptin-induced angiogenic and signaling responses were significantly inhibited with 100 nM Allo-aca. We also found that leptin increased its own mRNA and protein expression in both cell lines, and this autocrine effect was abolished by 100-250 nM Allo-aca.

Conclusions

Our data provide new insights into the role of leptin in ocular endothelial cells and represent the first original report on targeting ObR in ophthalmic cell models.     

Stress-induced expression and structure of the putative gene hyp-1 for hypericin biosynthesis

The phenolic oxidative coupling protein (Hyp-1) with proposed activity in the biosynthesis of hypericin in Hypericum perforatum shares about 50 % sequence similarity with Bet.v.1-like/PR-10 proteins. In our previous study, we showed that this protein is not a limiting factor in hypericin biosynthesis. To ascertain the role of Hyp-1 in defense mechanisms, we have analyzed some structural features of the hyp-1 gene in 14 Hypericum species with different abilities to synthesise hypericin. We show that the hyp-1 gene possesses characteristics typical for genes encoding plant PR-10 proteins. The coding sequence of the hyp-1 gene is interrupted by a single 86- to 125-bp intron localised strictly in codon 62, which is a typical feature of the dicot PR-10 subfamily. The localisation of the intron is conserved in all 14 tested Hypericum species indicating a common evolutionary history with genes encoding PR-10 proteins. In addition, we report that the hyp-1 gene exhibits a similar response to stress conditions as the PR-10 proteins encoding genes. Following either wounding or infection by Agrobacterium tumefaciens, all analysed Hypericum species exhibited rapid and significant upregulation of hyp-1 gene expression; this was particularly observed in hypericin-producing species. On the other hand, in the presence of high levels of abscisic acid, different levels of gene expression were observed.     

Agrobacterium bohemicum sp. nov. isolated from poppy seed wastes in central Bohemia

Two non-pathogenic strains R89-1 and R90^T isolated from poppy seed (Papaver somniferum L.) wastes were phenotypically and genotypically characterized. Multilocus sequence analysis (MLSA) was conducted with six genes (atpD, glnA, gyrB, recA, rpoB, 16S rRNA). The strains represented a new species which clustered with Agrobacterium rubi NBRC 13261^T and Agrobacterium skierniewicense Ch11^T type strains. MLSA was further accompanied by whole-genome phylogeny, in silico DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) analyses for both strains. ANI and dDDH values were deep below the species delineation threshold. Phenotypic features of the novel strains unequivocally allowed their differentiation from all other Agrobacterium species. Unlike other agrobacteria, the strains were salt sensitive and were able to biotransform morphine alkaloids. The dominant cellular fatty acids are 18:1 w7c, 16:0 and 12:0 aldehyde/16:1 iso I/14:0 3OH summed in feature 2 and the major respiratory quinine is Q-10 (87%). The DNA G + C content is 56 mol%. Microbial community analysis indicated probable association with P. somniferum plant material. Altogether, these characteristics showed that strains R90^T and R89-1 represent a new species of the genus Agrobacterium which we propose to name Agrobacterium bohemicum. The type strain of A. bohemicum is R90^T (=CCM 8736^T = DSM 104667^T).     

Targeted deletion of RIC8A in mouse neural precursor cells interferes with the development of the brain,eyes, and muscles

Autosomal recessive disorders such as Fukuyama congenital muscular dystrophy, Walker–Warburg syndrome, and the muscle–eye–brain disease are characterized by defects in the development of patient's brain, eyes, and skeletal muscles. These syndromes are accompanied by brain malformations like type II lissencephaly in the cerebral cortex with characteristic overmigrations of neurons through the breaches of the pial basement membrane. The signaling pathways activated by laminin receptors, dystroglycan and integrins, control the integrity of the basement membrane, and their malfunctioning may underlie the pathologies found in the rise of defects reminiscent of these syndromes. Similar defects in corticogenesis and neuromuscular disorders were found in mice when RIC8A was specifically removed from neural precursor cells. RIC8A regulates a subset of G‐protein α subunits and in several model organisms, it has been reported to participate in the control of cell division, signaling, and migration. Here, we studied the role of RIC8A in the development of the brain, muscles, and eyes of the neural precursor‐specific conditional Ric8a knockout mice. The absence of RIC8A severely affected the attachment and positioning of radial glial processes, Cajal‐Retzius’ cells, and the arachnoid trabeculae, and these mice displayed additional defects in the lens, skeletal muscles, and heart development. All the discovered defects might be linked to aberrancies in cell adhesion and migration, suggesting that RIC8A has a crucial role in the regulation of cell–extracellular matrix interactions and that its removal leads to the phenotype characteristic to type II lissencephaly‐associated diseases. © 2018 Wiley Periodicals, Inc. Develop Neurobiol 78: 374–390, 2018     

Lamb ductus arteriosus: Effect of prostaglandin synthesis inhibitors on the muscle tone and the response to prostaglandin E2

The prostaglandin synthesis inhibitors, indomethacin and eicosa-5,8,11, 14-tetraynoic acid (ETA), have been tested on the isolated lamb ductus arteriosus at low and high PO₂ levels. Both compounds produced a gradual contraction of the hypoxic vessel, and at equal doses the effect of indomethacin was stronger. The maximal tension output of the hypoxic tissue under indomethacin was equal to that of the oxygen-contracted control. ETA- and indomethacin-treated preparations contracted further upon transfer from a low to a high oxygen environment, and the response under indomethacin exceeded significantly control values. Control preparations were relaxed markedly by PGE₂ in low oxygen but showed little or no response in high oxygen. In contrast, preparations pretreated with the inhibitors retained their sensitivity to PGE₂ during exposure to high oxygen. The data are consistent with the idea that E-type prostaglandins play a role in the regulation of the intrinsic tone of the ductus arteriosus during foetal life. It is also suggested that the sensitivity of ductal muscle to E-type prostaglandins is controlled by the rate of endogenous prostaglandin formation.