Characterization of the Viable but Nonculturable (VBNC) State in Saccharomyces cerevisiae

The Viable But Non Culturable (VBNC) state has been thoroughly studied in bacteria. In contrast, it has received much less attention in other microorganisms. However, it has been suggested that various yeast species occurring in wine may enter in VBNC following sulfite stress.In order to provide conclusive evidences for the existence of a VBNC state in yeast, the ability of Saccharomyces cerevisiae to enter into a VBNC state by applying sulfite stress was investigated. Viable populations were monitored by flow cytometry while culturable populations were followed by plating on culture medium. Twenty-four hours after the application of the stress, the comparison between the culturable population and the viable population demonstrated the presence of viable cells that were non culturable. In addition, removal of the stress by increasing the pH of the medium at different time intervals into the VBNC state allowed the VBNC S. cerevisiae cells to “resuscitate”. The similarity between the cell cycle profiles of VBNC cells and cells exiting the VBNC state together with the generation rate of cells exiting VBNC state demonstrated the absence of cellular multiplication during the exit from the VBNC state. This provides evidence of a true VBNC state. To get further insight into the molecular mechanism pertaining to the VBNC state, we studied the involvement of the SSU1 gene, encoding a sulfite pump in S. cerevisiae. The physiological behavior of wild-type S. cerevisiae was compared to those of a recombinant strain overexpressing SSU1 and null Δssu1 mutant. Our results demonstrated that the SSU1 gene is only implicated in the first stages of sulfite resistance but not per se in the VBNC phenotype. Our study clearly demonstrated the existence of an SO₂-induced VBNC state in S. cerevisiae and that the stress removal allows the “resuscitation” of VBNC cells during the VBNC state.     

Discovery of cyclic sulfoxide hydroxyethylamines as potent and selective β-site APP-cleaving enzyme 1 (BACE1) inhibitors: Structure based design and in vivo reduction of amyloid β-peptides

Previous structure based optimization in our laboratories led to the identification of a novel, high-affinity cyclic sulfone hydroxyethylamine-derived inhibitor such as 1 that lowers CNS-derived Aβ following oral administration to transgenic APP51/16 mice. Herein we report SAR development in the S3 and S2′ subsites of BACE1 for cyclic sulfoxide hydroxyethyl amine inhibitors, the synthetic approaches employed in this effort, and in vivo data for optimized compound such as 11d.     

Focused library development of 2-phenylacrylamides as broad spectrum cytotoxic agents

With our lead compound (E)-3-(4-chlorophenyl)-2-(1H-pyrrole-2-carbonyl)acrylonitrile (1) inducing 50% growth inhibition of 11 cancer cell lines at 27–61 μM, potency enhancements were rapidly established through the synthesis of a series of focused compound libraries. Six highly focused libraries (46 compounds in total) were synthesised. Each library allowed the identification of a new lead compound, viz Library A identified (E)-3-(pentafluorophenyl)-2-(1H-pyrrole-2-carbonyl)acrylonitrile (11) and (E)-3-(1H-indol-3-yl)-2-(1H-pyrrole-2-carbonyl)acrylonitrile (13) as inhibitors with improved cytotoxicity. Synthesis of discrete libraries of amidoacrylamide analogues (Ar–CC(CN)–Ar?Ar–CC(CN)–C(O)NH)–Ar) resulted in a series of analogues significantly more potent that the lead, 1. Three furan three analogues: (E)-3-(5-chlorofuran-2-yl)-2-cyano-N-(4-methoxybenzyl)acrylamide (33), (E)-3-(5-bromofuran-2-yl)-2-cyano-N-(4-methoxybenzyl)acrylamide (34) and (E)-2-cyano-3-(furan-3-yl)-N-(4-methoxybenzyl)acrylamide (37) returned broad spectrum growth inhibition (GI₅₀ values of 5–16 μM). Replacement of the furan moiety with simple aromatics gave an additional three analogues: (E)-2-cyano-N-(4-methoxybenzyl)-3-phenylacrylamide (39), (E)-3-(4-chlorophenyl)-2-cyano-N-(4-methoxybenzyl)acrylamide (41) and (E)-2-cyano-N-(4-methoxyphenyl)-3-(naphthalen-1-yl)acrylamide (45) with GI₅₀ values of 7–24 μM. The final library retained the aromatic substituents but introduced a 3,4-dichlorbenzylamine moiety to afford the 1-naphthyl substituted 52, which was the most potent broad spectrum cytotoxic analogue produced here in with an average GI₅₀ = 8.6 μM. This represents a fivefold potency enhancement relative to 1 and a new cytotoxic scaffold suitable for further development.     

Soil chemistry,elemental profiles and elemental distribution in nickel hyperaccumulator species from New Caledonia

Plant and Soil - This study aimed to establish elemental profiles and to spatially resolve the elemental distribution in five New Caledonian woody Ni hyperaccumulator plant species (Geissois...     

Discovering new clinical markers in the field of glycomics

Glycosylation modifications have been reported in a number of disease states and, as a result, there is significant focus on the discovery and development of glycan-based biomarkers. Glyco-biomarkers have the potential to enhance the efficacy and efficiency of the diagnostic procedures for these diseases.     

Modulation of Na(+)-K(+)-ATPase cell surface abundance through structural determinants on the α1-subunit

Through their ion-pumping and non-ion-pumping functions, Na(+)-K(+)-ATPase protein complexes at the plasma membrane are critical to intracellular homeostasis and to the physiological and pharmacological actions of cardiotonic steroids. Alteration of the abundance of Na(+)-K(+)-ATPase units at the cell surface is one of the mechanisms for Na(+)-K(+)-ATPase regulation in health and diseases that has been closely examined over the past few decades. We here summarize these findings, with emphasis on studies that explicitly tested the involvement of defined regions or residues on the Na(+)-K(+)-ATPase α1 polypeptide. We also report new findings on the effect of manipulating Na(+)-K(+)-ATPase membrane abundance by targeting one of these defined regions: a dileucine motif of the form [D/E]XXXL[L/I]. In this study, opossum kidney cells stably expressing rat α1 Na(+)-K(+)-ATPase or a mutant where the motif was disrupted (α1-L499V) were exposed to 30 min of substrate/coverslip-induced-ischemia followed by reperfusion (I-R). Biotinylation studies suggested that I-R itself acted as an inducer of Na(+)-K(+)-ATPase internalization and that surface expression of the mutant was higher than the native Na(+)-K(+)-ATPase before and after ischemia. Annexin V/propidium iodide staining and lactate dehydrogenase release suggested that I-R injury was reduced in α1-L499V-expressing cells compared with α1-expressing cells. Hence, modulation of Na(+)-K(+)-ATPase cell surface abundance through structural determinants on the α-subunit is an important mechanism of regulation of cellular Na(+)-K(+)-ATPase in various physiological and pathophysiological conditions, with a significant impact on cell survival in face of an ischemic stress.