Cellular transport and homeostasis of essential and nonessential metals

Metals can have a number of detrimental or beneficial effects in the cell, but first they must get in. Organisms have evolved transport mechanisms to get metals that are required, or essential into the cell. Nonessential metals often enter the cell through use of the machinery provided for essential metals. Much work has been done to advance our understanding of how these metals are transported across plasma and organelle membranes. This review provides an overview of essential and nonessential metal transport and homeostatic processes.     

Akt3 deficiency in macrophages promotes foam cell formation and atherosclerosis in mice

Akt, a serine-threonine protein kinase, exists as three isoforms. The Akt signaling pathway controls multiple cellular functions in the cardiovascular system, and the atheroprotective endothelial cell-dependent role of Akt1 has been recently demonstrated. The role of Akt3 isoform in cardiovascular pathophysiology is not known. We explored the role of Akt3 in atherosclerosis using mice with a genetic ablation of the Akt3 gene. Using hyperlipidemic ApoE(-/-) mice, we demonstrated a macrophage-dependent, atheroprotective role for Akt3. In vitro experiments demonstrated differential subcellular localization of Akt1 and Akt3 in macrophages and showed that Akt3 specifically inhibits macrophage cholesteryl ester accumulation and foam cell formation, a critical early event in atherogenesis. Mechanistically, Akt3 suppresses foam cell formation by reducing lipoprotein uptake and promoting ACAT-1 degradation via the ubiquitin-proteasome pathway. These studies demonstrate the nonredundant atheroprotective role for Akt3 exerted via the previously unknown link between the Akt signaling pathway and cholesterol metabolism.     

Effect of loop length variation on quadruplex-Watson Crick duplex competition

The effect of loop length on quadruplex stability has been studied when the G-rich strand is present along with its complementary C-rich strand, thereby resulting in competition between quadruplex and duplex structures. Using model sequences with loop lengths varying from T to T5, we carried out extensive FRET to discover the influence of loop length on the quadruplex-Watson Crick duplex competition. The binding data show an increase in the binding affinity of quadruplexes towards their complementary strands upon increasing the loop length. Our kinetic data reveal that unfolding of the quadruplex in presence of a complementary strand involves a contribution from a predominant slow and a small population of fast opening conformer. The contribution from the fast opening conformer increases upon increasing the loop length leading to faster duplex formation. FCS data show an increase in the interconversion between the quadruplex conformers in presence of the complementary strand, which shifts the equilibrium towards the fast opening conformer with an increase in loop length. The relative free-energy difference (Delta DeltaG(o)) between the duplex and quadruplex indicates that an increase in loop length favors duplex formation and out competes the quadruplex.     

Sustaining immunity during starvation in bivalve mollusc: A costly affair

Complete or partial depletion of resource in a freshwater habitat is a common phenomenon. As a consequence, aquatic fauna including bivalve molluscs may be exposed to dietary stress on a seasonal basis. Haemocyte based innate immune profile of the freshwater mollusc Lamellidens marginalis (Bivalvia: Eulamellibranchiata) was evaluated under starvation induced stress for a maximum period of 32 days in a controlled laboratory condition. During starvation, the bivalve haemocytes maintained a homeostasis in phagocytic efficacy and nitric oxide generation ability with respect to the control. The mollusc maintained a significantly high protein content in its haemolymph and tissues under the nutritional stress with respect to the control. The dietary stress had no significant impact on the activity of digestive tissue derived α-amylase till sixteenth day but by 32 days the enzyme activity went down significantly. The histopathological profile revealed that the bivalve was adapted to maintain a steady immune profile by incurring degeneration of its own tissue structure. The total haemocyte count surged significantly till 16 days but differed insignificantly with respect to the control at 32 days implying probable haematopoietic exhaustion. The study reflects the instinctive urge of the bivalve to maintain immune physiology at heavy metabolic cost under nutrient limited condition.     

Enhanced adsorption of congo red from aqueous solutions by chitosan hydrogel beads impregnated with cetyl trimethyl ammonium bromide

The adsorption of congo red (CR) onto chitosan (CS) beads impregnated by a cationic surfactant (CTAB, cetyl trimethyl ammonium bromide) was investigated. Chitosan beads impregnated at a ratio of 1/20 of CTAB to CS (0.05% of CTAB and 1% of CS) increased the CR adsorption capacity by 2.2 times from 162.3 mg/g (0% CTAB) to 352.5 mg/g (0.05% CTAB). The CR adsorption decreased with an increase in pH of the CR solution from 4.0 to 9.0. The Sips isotherm model showed a good fit with the equilibrium experimental data and the values of the heterogeneity factor (n) indicated heterogeneous adsorption of CR onto CS/CTAB beads, as well as CS beads. The kinetic data showed better fit to the pseudo second-order rate model than to the pseudo first-order rate model. The impregnation of CS beads by cationic surfactants showed the highest adsorption capacities of CR compared to any other adsorbents and would be a good method to increase adsorption efficiency for the removal of anionic dyes in a wastewater treatment process.     

ortho-Carom-N bond fusion in aniline associated with electrophilic chlorination reactions at ruthenium(III) coordinated acetylacetonates

In an unusual reaction of [Ru^III(acac)₂(CH₃CN)₂](ClO₄) ([1], acac = acetylacetonate) and aniline (Ph-NH₂), resulted in the formation of ortho-semidine due to dimerisation of aniline via oxidative ortho-C_arom-N bond formation reaction. This oxidation reaction is associated with stepwise chlorination of coordinated acac ligands at the γ-carbon atom resulting in the formation of [Ru^III(acac)₂L⁻] [2a], [Ru^III(Cl-acac)(acac)L⁻] [2b], [Ru^III(acac)(Cl-acac)L⁻] [2c] and [Ru^III(Cl-acac)₂L⁻] [2d] (L⁻ = N-phenyl-ortho-semiquinonediimine) complexes, respectively. These have been characterized by ¹H NMR, UV-Vis-NIR, ESI-MS and cyclic voltammetry studies. Single crystal X-ray structures of 2c and 2d are reported. Crystallographic structural bond parameters of 2c and 2d revealed bond length equalization of C-C, C-O and M-O bonds. It has been shown that perchlorate () counter anion, present in the starting ruthenium complex, acts as the oxidizing agent in bringing about oxidation of Ph-NH₂ to ortho-semidine. The chloronium ions, produced in situ, chlorinate the coordinated acac ligands at the γ-carbon atom. Such electrophilic substitution of coordinated acac ligands indicates that the Ru-acac metallacycles in the reference compounds are aromatic. The complexes showed an intense and featureless band centered near 520 nm, and a structured band near 275 nm. These displayed one reversible cathodic response in the range, −1.1 to −0.8 V and one reversible anodic response between 0.4 and 0.6 V versus the Saturated Calomel reference Electrode, SCE. The response at the anodic potential is due to oxidation of the coordinated ligand L, while the reversible response at cathodic potential is due to reduction of the metal center.     

LNA mediated in situ hybridization of miR171 and miR397a in leaf and ambient root tissues revealed expressional homogeneity in response to shoot heat shock in Arabidopsis thaliana

MicroRNAs (miRNAs) are endogenous non-protein coding RNA molecules of approximately 21 nucleotides in length capable of modulating gene expression in animals and plants. The role of miRNA based gene regulation has been proved in several pathways including in plant growth, development and stress response. In this study miR171 and miR397a were tested for their expression pattern under different heat shock regimes in shoot and root tissues of Arabidopsis thaliana using Locked Nucleic Acid (LNA) mediated in situ hybridization. With an increase in temperature across 35 °C, 40 °C and 45 °C there was a corresponding increased up-regulation of miR171 in leaf tissues compared to ambient temperature. Similarly, an unambiguous elevated expression of miR171 within increase in duration of exposure at each temperature regime across 1 h, 2 h and 3 h was noticed in comparison to ambient control leaf tissue. On the other hand, miR397a, which expressed at ambient control conditions, got down-regulated both with increase in heat and exposure regime in leaf tissues. Both miRNAs expressed in control ambient root tissues. Maintaining the root zone temperature at ambient conditions, upon imposing heat shock regime to shoot system, miR171 recorded corresponding increased up-regulation as indicated by the intensity of in situ hybridization, while miR397a got down-regulated. Given the differential homogeneity in expression pattern of both miRNA in leaf and root tissues experiencing heat shock regimes, possibilities of movement of heat shock induced signals to root tissues seem to be obvious.     

Phosphoproteomic Analysis of Platelets Activated by Pro-Thrombotic Oxidized Phospholipids and Thrombin

Specific oxidized phospholipids (oxPC_CD36) promote platelet hyper-reactivity and thrombosis in hyperlipidemia via the scavenger receptor CD36, however the signaling pathway(s) induced in platelets by oxPC_CD36 are not well defined. We have employed mass spectrometry-based tyrosine, serine, and threonine phosphoproteomics for the unbiased analysis of platelet signaling pathways induced by oxPC_CD36 as well as by the strong physiological agonist thrombin. oxPC_CD36 and thrombin induced differential phosphorylation of 115 proteins (162 phosphorylation sites) and 181 proteins (334 phosphorylation sites) respectively. Most of the phosphoproteome changes induced by either agonist have never been reported in platelets; thus they provide candidates in the study of platelet signaling. Bioinformatic analyses of protein phosphorylation dependent responses were used to categorize preferential motifs for (de)phosphorylation, predict pathways and kinase activity, and construct a phosphoproteome network regulating integrin activation. A putative signaling pathway involving Src-family kinases, SYK, and PLCγ2 was identified in platelets activated by oxPC_CD36. Subsequent ex vivo studies in human platelets demonstrated that this pathway is downstream of the scavenger receptor CD36 and is critical for platelet activation by oxPC_CD36. Our results provide multiple insights into the mechanism of platelet activation and specifically in platelet regulation by oxPC_CD36.