Differential responses to salinity of two Atriplex halimus populations in relation to organic solutes and antioxidant systems involving thiol reductases

Atriplex halimus L. is a xero-halophyte species widespread in the Mediterranean basin. The tolerance to water stress and high salinity of two Atriplex populations from semi-arid (Djelfa) and arid saline (Laghouat) Algerian regions has been investigated in relation with organic solutes and antioxidant systems. Whereas no noticeable difference was observed between the two populations under water stress resulting from withholding watering or PEG treatment, Laghouat plants display significantly higher fresh and dry weights than Djelfa plants when exposed to high salinity. At 300mM NaCl, Laghouat plants exhibit higher concentrations in Na(+), proline and quaternary ammonium compounds, and a higher catalase activity than Djelfa plants. We then analysed the involvement of recently characterized plastidial thiol reductases, peroxiredoxins (Prxs) and methionine sulphoxide reductases (MSRs), key enzymes scavenging organic peroxides and repairing oxidized proteins, respectively. Upon salt treatment (300mM NaCl), we observed higher amounts of PrxQ and over-oxidized 2-Cys Prx in Laghouat than in Djelfa. An increased abundance of plastidial MSRA and a higher total MSR activity were also noticed in Laghouat plants treated with 300mM NaCl compared to Djelfa ones. We propose that mechanisms based on organic solutes and antioxidant enzymes like catalases, peroxiredoxins and MSRs party underlie the better tolerance of the Laghouat population to high salt.     

Ultimate Use of Two-Photon Fluorescence Microscopy to Map Orientational Behavior of Fluorophores

The orientational distribution of fluorophores is an important reporter of the structure and function of their molecular environment. Although this distribution affects the fluorescence signal under polarized-light excitation, its retrieval is limited to a small number of parameters. Because of this limitation, the need for a geometrical model (cone, Gaussian, etc.) to effect such retrieval is often invoked. In this work, using a symmetry decomposition of the distribution function of the fluorescent molecules, we show that polarized two-photon fluorescence based on tunable linear dichroism allows for the retrieval of this distribution with reasonable fidelity and without invoking either an a priori knowledge of the system to be investigated or a geometrical model. We establish the optimal level of detail to which any distribution can be retrieved using this technique. As applied to artificial lipid vesicles and cell membranes, the ability of this method to identify and quantify specific structural properties that complement the more traditional molecular-order information is demonstrated. In particular, we analyze situations that give access to the sharpness of the angular constraint, and to the evidence of an isotropic population of fluorophores within the focal volume encompassing the membrane. Moreover, this technique has the potential to address complex situations such as the distribution of a tethered membrane protein label in an ordered environment.     

Carbonic anhydrase inhibitors: 2-substituted-1,3,4-thiadiazole-5-sulfamides act as powerful and selective inhibitors of the mitochondrial isozymes VA and VB over the cytosolic and membrane-associated carbonic anhydrases I, II and IV

A series of 2-substituted-1,3,4-thiadiazole-5-sulfamides was prepared and assayed as inhibitors of several carbonic anhydrase (CA, EC 4.2.1.1) isoforms, the cytosolic CA I and II, the membrane-associated CA IV and the mitochondrial CA VA and VB. The new compounds showed weak inhibitory activity against hCA I (K(I)s of 102 nM-7.42 microM), hCA II (K(I)s of 0.54-7.42 microM) and hCA IV (K(I)s of 4.32-10.05 microM) but were low nanomolar inhibitors of hCA VA and hCA VB, with inhibition constants in the range of 4.2-32 nM and 1.3-74 nM, respectively. Furthermore, the selectivity ratios for inhibiting the mitochondrial enzymes over CA II were in the range of 67.5-415, making these sulfamides the first selective CA VA/VB inhibitors.     

Carbonic anhydrase inhibitors: Selective inhibition of the extracellular, tumor-associated isoforms IX and XII over isozymes I and II with glycosyl-thioureido-sulfonamides

A series of glycosyl-thioureido sulfonamides incorporating glucosamine, galactosamine, and mannosamine tails, and sulfanilamide, halogenosulfanilamide, and metanilamide heads was synthesized. Many of the new compounds showed micromolar-submicromolar affinity for the inhibition of the cytosolic isoforms I and II of the metalloenzyme carbonic anhydrase (CA, EC 4.2.1.1), but low nanomolar binding to the tumor-associated isozymes, CA IX and XII. The selectivity ratios for the inhibition of the tumor-associated over the cytosolic isozymes were in the range of 107-955 for the most selective such inhibitors.     

Ultimate Use of Two-Photon Fluorescence Microscopy to Map Orientational Behavior of Fluorophores

The orientational distribution of fluorophores is an important reporter of the structure and function of their molecular environment. Although this distribution affects the fluorescence signal under polarized-light excitation, its retrieval is limited to a small number of parameters. Because of this limitation, the need for a geometrical model (cone, Gaussian, etc.) to effect such retrieval is often invoked. In this work, using a symmetry decomposition of the distribution function of the fluorescent molecules, we show that polarized two-photon fluorescence based on tunable linear dichroism allows for the retrieval of this distribution with reasonable fidelity and without invoking either an a priori knowledge of the system to be investigated or a geometrical model. We establish the optimal level of detail to which any distribution can be retrieved using this technique. As applied to artificial lipid vesicles and cell membranes, the ability of this method to identify and quantify specific structural properties that complement the more traditional molecular-order information is demonstrated. In particular, we analyze situations that give access to the sharpness of the angular constraint, and to the evidence of an isotropic population of fluorophores within the focal volume encompassing the membrane. Moreover, this technique has the potential to address complex situations such as the distribution of a tethered membrane protein label in an ordered environment.