Interactions between permeation and gating in the TMEM16B/anoctamin2 calcium-activated chloride channel

At least two members of the TMEM16/anoctamin family, TMEM16A (also known as anoctamin1) and TMEM16B (also known as anoctamin2), encode Ca²⁺-activated Cl⁻ channels (CaCCs), which are found in various cell types and mediate numerous physiological functions. Here, we used whole-cell and excised inside-out patch-clamp to investigate the relationship between anion permeation and gating, two processes typically viewed as independent, in TMEM16B expressed in HEK 293T cells. The permeability ratio sequence determined by substituting Cl⁻ with other anions (P_X/P_Cl) was SCN⁻ > I⁻ > NO₃⁻ > Br⁻ > Cl⁻ > F⁻ > gluconate. When external Cl⁻ was substituted with other anions, TMEM16B activation and deactivation kinetics at 0.5 µM Ca²⁺ were modified according to the sequence of permeability ratios, with anions more permeant than Cl⁻ slowing both activation and deactivation and anions less permeant than Cl⁻ accelerating them. Moreover, replacement of external Cl⁻ with gluconate, or sucrose, shifted the voltage dependence of steady-state activation (G-V relation) to more positive potentials, whereas substitution of extracellular or intracellular Cl⁻ with SCN⁻ shifted G-V to more negative potentials. Dose–response relationships for Ca²⁺ in the presence of different extracellular anions indicated that the apparent affinity for Ca²⁺ at +100 mV increased with increasing permeability ratio. The apparent affinity for Ca²⁺ in the presence of intracellular SCN⁻ also increased compared with that in Cl⁻. Our results provide the first evidence that TMEM16B gating is modulated by permeant anions and provide the basis for future studies aimed at identifying the molecular determinants of TMEM16B ion selectivity and gating.     

Interaction of metallothionein with tumor suppressor p53 protein

Previous reports have shown that metallothionein (MT) may modulate p53 activity through zinc exchange. However, little is known on a direct interaction between MT and p53 in cells. The results demonstrate an interaction between MT and p53 can occur in vitro. The complex between MT and p53 was observed in breast cancer epithelial cells with both wild and inactive type of p53. Furthermore, it was shown that wt-p53 was preferentially associated with Apo-MT. Our data suggest that co-expression of MT and p53 and their complex formation in tumor cells may be involved in regulation of apoptosis in these cells.     

NMR analysis of Caenorhabditis elegans FLP-18 neuropeptides: implications for NPR-1 activation

Phe-Met-Arg-Phe-NH2 (FMRFamide)-like peptides (FLPs) are the largest neuropeptide family in animals, particularly invertebrates. FLPs are characterized by a C-N-terminal gradient of decreasing amino acid conservation. Neuropeptide receptor 1 (NPR-1) is a G-protein coupled receptor (GPCR), which has been shown to be a strong regulator of foraging behavior and aggregation responses in Caenorhabditis elegans. Recently, ligands for NPR-1 were identified as neuropeptides coded by the precursor genes flp-18 and flp-21 in C. elegans. The flp-18 gene encodes eight FLPs including DFDGAMPGVLRF-NH2 and EMPGVLRF-NH2. These peptides exhibit considerably different activities on NPR-1, with the longer one showing a lower potency. We have used nuclear magnetic resonance and biological activity to investigate structural features that may explain these activity differences. Our data demonstrate that long-range electrostatic interactions exist between N-terminal aspartates and the C-terminal penultimate arginine as well as N-terminal hydrogen-bonding interactions that form transient loops within DFDGAMPGVLRF-NH2. We hypothesize that these loops, along with peptide charge, diminish the activity of this peptide on NPR-1 relative to that of EMPGVLRF-NH2. These results provide some insight into the large amino acid diversity in FLPs.     

Ascorbic acid and EDTA treatment of lead toxicity in rats.

Ascorbic acid given orally to lead exposed rats has similar chelating properties as equimolar amounts of parenterally administered EDTA. Ascorbic acid in combination with EDTA is more than twice as effective as either chelating agent given alone and the combination is particularly effective in removing lead from the central nervous system.     

Transgenic plants with improved dehydration-stress tolerance: Progress and future prospects

This review summarizes the recent progress made towards the development of transgenic plants with improved tolerance to water stress and salinity. Of the various strategies employed, emphasis has been given to the genes engineered for the biosynthesis of osmoprotectants and osmolytes. This review also briefly discusses the importance of the use of specific stress inducible promoters and the future prospects of transgenic plants with improved agronomic traits.