Structure and property based design of factor Xa inhibitors: pyrrolidin-2-ones with monoaryl P4 motifs

Structure and property based drug design was exploited in the synthesis of sulfonamidopyrrolidin-2-one-based factor Xa inhibitors, incorporating neutral and basic monoaryl P4 groups, ultimately producing potent inhibitors with effective levels of anticoagulant activity and extended oral pharmacokinetic profiles. However, time dependant inhibition of Cytochrome P450 3A4 was a particular issue with this series.     

The MIT user center for neutron capture therapy research

Neutron capture therapy (NCT) research encompasses a wide range of preclinical and clinical studies needed to develop this promising but complex cancer treatment. Many specialized facilities and capabilities including thermal and epithermal neutron irradiation facilities, boron analysis, specialized mixed-field dosimetry, animal care facilities and protocols, cell culture laboratories, and, for human clinical studies, licenses and review board approvals are required for NCT research. Such infrastructure is essential, but much of it is not readily available within the community. This is especially true for neutron irradiation facilities, which often require significant development and capital investment too expensive to duplicate at each site performing NCT research. To meet this need, the NCT group at the Massachusetts Institute of Technology (MIT) has established a User Center for NCT researchers that is already being accessed successfully by various groups. This paper describes the facilities, capabilities and other resources available at MIT and how the NCT research community can access them.     

Survival and epiphytic fitness of a nonpathogenic mutant of Xanthomonas campestris pv. glycines

Xanthomonas campestris pv. glycines is the causal agent of bacterial pustule disease of soybeans. The objective of this work was to construct a nonpathogenic mutant derived from the pathogenic wild-type strain YR32 and to evaluate its effectiveness in preventing growth of its parent on the soybean phyllosphere. A mini-Tn5-derived transposon was used to generate nonpathogenic mutants. Southern hybridization and pulsed-field gel electrophoresis confirmed the presence of a single transposon in each of the nonpathogenic mutants. One of the nonpathogenic mutants, M715, failed to induce a hypersensitive response in tomato leaves. An ice nucleation gene (inaZ) carried in pJL1703 was introduced into strain YR32 as a reporter gene to demonstrate that the presence of M715 could reduce colonization of the soybean phyllosphere by YR32. de Wit serial replacement analysis showed that M715 competed equally with its wild-type parental strain, YR32. Epiphytic fitness analysis of YR32 in the greenhouse indicated that the population dynamics of strains YR32, YR32(pJL1703), and M715 were similar, although the density of the mutant was slightly less than that of its parent. The M715 mutant was able to survive for 16 days after inoculation on soybean leaves and maintained population densities of approximately 10(4) to 10(5) cells g (fresh weight) of leaf(-1). Therefore, M715 shows promise as an effective biocontrol agent for bacterial pustule disease in soybeans.     

<Emphasis Type="Italic">Muscadinia rotundifolia</Emphasis> ‘Noble’ defense response to <Emphasis Type="Italic">Plasmopara viticola</Emphasis> inoculation by inducing phytohormone-mediated stilbene accumulation

Downy mildew (DM), one of the most devastating grape diseases worldwide, is caused by the biotrophic oomycete Plasmopara viticola (Pv). In general, grapevine responds to Pv infection with the accumulation of phytoalexins as part of the innate immune system, and diverse phytoalexins are induced on grapevines with different DM-resistance levels in response to Pv invasion. However, the regulation of phytoalexin biosynthesis during grapevine against Pv is still unclear. Herein, we detected stilbenes by UPLC-ESI-MS/MS and found that resveratrol was accumulated to higher level and earlier in the DM-immune Muscadinia rotundifolia ‘Noble’ than that in the DM-susceptible Vitis vinifera ‘Thompson Seedless’ after Pv inoculation. Additionally, a considerable amount of pterostilbene and ε-viniferin was found in ‘Noble’, while a little was detected in ‘Thompson Seedless’. Resveratrol was glycosylated into piceid both in ‘Noble’ and ‘Thompson Seedless’ after Pv inoculation. The qPCR analysis of gene expression indicated that the resveratrol-synthesis gene (STS) was induced by Pv inoculation earlier in ‘Noble’ than that in ‘Thompson Seedless’, while the pterostilbene-synthesis gene (ROMT) was induced in ‘Noble’ but not in ‘Thompson Seedless’ at all. The piceid-synthesis gene (GT) was generally up-regulated in both cultivars. Sequence analysis of STS, ROMT, and GT promoters revealed that they contained cis-regulatory elements responsive to phytohormones and pathogens. Following Pv inoculation, the level of SA, MeJA, and ABA was found to be consistently higher in ‘Noble’ than those in ‘Thompson Seedless’. The results of exogenous hormone elicitation further demonstrated that the accumulation of stilbenes was regulated by phytohormones. The earlier and higher accumulation of phytohormones and consequent induction of stilbene synthesis may play an important role in grapevine defense against downy mildew disease.     

Actin filaments regulate the adhesion between the plasma membrane and the cell wall of tobacco guard cells

During the opening and closing of stomata, guard cells undergo rapid and reversible changes in their volume and shape, which affects the adhesion of the plasma membrane (PM) to the cell wall (CW). The dynamics of actin filaments in guard cells are involved in stomatal movement by regulating structural changes and intracellular signaling. However, it is unclear whether actin dynamics regulate the adhesion of the PM to the CW. In this study, we investigated the relationship between actin dynamics and PM–CW adhesion by the hyperosmotic-induced plasmolysis of tobacco guard cells. We found that actin filaments in guard cells were depolymerized during mannitol-induced plasmolysis. The inhibition of actin dynamics by treatment with latrunculin B or jasplakinolide and the disruption of the adhesion between the PM and the CW by treatment with RGDS peptide (Arg-Gly-Asp-Ser) enhanced guard cell plasmolysis. However, treatment with latrunculin B alleviated the RGDS peptide-induced plasmolysis and endocytosis. Our results reveal that the actin depolymerization is involved in the regulation of the PW–CW adhesion during hyperosmotic-induced plasmolysis in tobacco guard cells.     

Overexpression of <Emphasis Type="Italic">AlTMP2</Emphasis> gene from the halophyte grass <Emphasis Type="Italic">Aeluropus littoralis</Emphasis> in transgenic tobacco enhances tolerance to different abiotic stresses by improving membrane stability and deregulating some stress-related genes

Herein, we report isolation of the AlTMP2 gene from the halophytic C4 grass Aeluropus littoralis. The subcellular localization suggested that AlTMP2 is a plasma membrane protein. In A. littoralis exposed to salt and osmotic stresses, the AlTMP2 gene was induced early and at a high rate, but was upregulated relatively later in response to abscisic acid and cold treatments. Expression of AlTMP2 in tobacco conferred improved tolerance against salinity, osmotic, H₂O₂, heat, and freezing stresses at the germination and seedling stages. Under control conditions, no growth or yield penalty were mentioned in transgenic plants due to the constitutive expression of AlTMP2. Interestingly, under greenhouse conditions, the seed yield of transgenic plants was significantly higher than that of non-transgenic (NT) plants grown under salt or drought stress. Furthermore, AlTMP2 plants had less electrolyte leakage, higher membrane stability, and lower Na⁺ and higher K⁺ accumulation than NT plants. Finally, six stress-related genes were shown to be deregulated in AlTMP2 plants relative to NT plants under both control and stress conditions. Collectively, these results indicate that AlTMP2 confers abiotic stress tolerance by improving ion homeostasis and membrane integrity, and by deregulating certain stress-related genes.     

Nectar and oleiferous trichomes as floral attractants in <Emphasis Type="Italic">Bulbophyllum saltatorium</Emphasis> Lindl. (Orchidaceae)

Although many Orchidaceae have deceit flowers that produce no reward, the most common reward, when present, is nectar. Bulbophyllum, however, is unusual in that the labellar secretions of most species investigated to date lack sugars, and, therefore, cannot be considered true nectar. The African species Bulbophyllum saltatorium is an exception in that it produces not only nectar but also possesses specialized, capitate oleiferous trichomes. The nectary of B. saltatorium is borne on the labellum and is represented by a deep, narrow, median longitudinal groove, having a small aperture, and flanked by trichomes. Isodiametric epidermal cells lining this groove secrete nectar which collects both in the groove and on the surface of the labellum. As well as a nectary, the labellum of B. saltatorium also bears three types of unicellular trichomes: the longest trichomes are borne distally and abaxially; the marginal ones form a rim around the entire labellum, and finally, massive, capitate trichomes occur proximally and adaxially. These are oleiferous, containing large quantities of oil which might function as precursors of volatile components of fragrance or provide a food-reward. To the best of our knowledge, this is the first time for such oleiferous trichomes to be described for Bulbophyllum. Therefore, apart from their color and markings, flowers of this species are able to attract pollinators in at least two, possibly three ways: food-reward in the form of nectar; fragrance; and possibly food-rewards in the form of food-hairs.     

Effect of mercury on pollen germination and tube growth in <Emphasis Type="Italic">Lilium longiflorum</Emphasis>

Pollen development and germination were adversely affected by the presence of mercury, whereas low-concentrations stimulated the whole procedure. Mercury caused morphological anomalies during the tube growth, characterized by irregularly increasing diameters and swelling tips. The main effect was the anomalous cell wall formation at the tip where a substantial number of organelles were found reducing the secretory vesicles. The dense organelle concentration caused a significant reduction of cytoplasmic movement integrity, and the cytosol streaming was gradually reduced or stopped completely. Electron dense, multilamellar myelin-like structures (MMS) of membranous material were frequently present, in close contact with plasmalemma or away from it. A loose network of fibrillar material and spherical aggregates mostly at the tip region were observed which progressively were loosened into the surrounding medium. Elevated mercury concentrations can affect plant reproduction, resulting in anomalies in gamete development and consequently loss of plant biodiversity.