UDP-N-Acetylglucosamine 2-Epimerase/N-Acetylmannosamine Kinase (GNE) Binds to Alpha-Actinin 1: Novel Pathways in Skeletal Muscle?

Hereditary inclusion body myopathy (HIBM) is a rare neuromuscular disorder caused by mutations in GNE, the key enzyme in the biosynthetic pathway of sialic acid. While the mechanism leading from GNE mutations to the HIBM phenotype is not yet understood, we searched for proteins potentially interacting with GNE, which could give some insights about novel putative biological functions of GNE in muscle. We used a Surface Plasmon Resonance (SPR)-Biosensor based assay to search for potential GNE interactors in anion exchanged fractions of human skeletal muscle primary culture cell lysate. Analysis of the positive fractions by in vitro binding assay revealed alpha-actinin 1 as a potential interactor of GNE. The direct interaction of the two proteins was assessed in vitro by SPR-Biosensor based kinetics analysis and in a cellular environment by a co-immunoprecipitation assay in GNE overexpressing 293T cells. Furthermore, immunohistochemistry on stretched mouse muscle suggest that both GNE and alpha-actinin 1 localize to an overlapping but not identical region of the myofibrillar apparatus centered on the Z line. The interaction of GNE with alpha-actinin 1 might point to its involvement in alpha-actinin mediated processes. In addition these studies illustrate for the first time the expression of the non-muscle form of alpha-actinin, alpha-actinin 1, in mature skeletal muscle tissue, opening novel avenues for its specific function in the sarcomere. Although no significant difference could be detected in the binding kinetics of alpha-actinin 1 with either wild type or mutant GNE in our SPR biosensor based analysis, further investigation is needed to determine whether and how the interaction of GNE with alpha-actinin 1 in skeletal muscle is relevant to the putative muscle-specific function of alpha-actinin 1, and to the muscle-restricted pathology of HIBM

Effect of Paraquat on Dark-Grown<i>Phaseolus vulgaris</i>Cells

Paraquat is known to affect all green plants and other eukaryotic organisms including mammalian cells. The aim of this study was to improve the understanding of paraquat toxicity in nonphotosynthetic plant cells using dark-grown kidney bean cells in tissue culture. It is shown that uptake of paraquat is an active process, and that paraquat inhibits cell growth, reduces DNA synthesis, and inhibits the activity of hydroxypyruvate reductase while enhancing the activity of glutathione reductase which is involved in cellular defense against oxidant stress. Additionally, it is demonstrated that iron ions are involved in paraquat toxicity. We conclude that uptake of paraquat into cells is via polyamine channels and that the deleterious effects of paraquat on these nonphotosynthetic cells are mediated by iron.</jats:p

Does metabolic water control the isotopic composition of water in microbial cells?

&amp;lt;p&amp;gt;Metabolic water, the water that is produced from O&amp;lt;sub&amp;gt;2&amp;lt;/sub&amp;gt; during respiration, carries an isotopic signature that can be different from that of the water the cell is growing in. It has been well known that for large land organisms, like birds and mammals, metabolic water contributes significantly to the water balance and has an important control on the signature of the oxygen-stable-isotopes of the water inside the organism. This isotopic signature is then carried over through isotopic equilibrium to other oxygen-bearing species like phosphate. However, for small organisms like bacteria, it has been widely assumed for decades, that the large surface area to volume ratio enables a fast exchange of the cell water with the ambient water. As a result, the isotopic signature of the metabolic water will be heavily diluted and erased. In contrast, a recent work reported indirect evidence of significant control of metabolic water on the oxygen isotopes inside microbial cells. This indirect evidence is based on deviations of oxygen isotopes in phosphate from the expected equilibrium with the ambient water. Here we report the results of experiments that directly tested the possible contribution of metabolic water to phosphate oxygen isotopes in bacteria. We found that ambient water did control the oxygen isotopes in the phosphate. However, there were large deviations from the expected equilibrium. Nevertheless, we found that these deviations were not correlated with the isotopic composition of metabolic water. Hence, other mechanisms, which will be discussed, are responsible for these deviations.&amp;lt;/p&amp;gt;</jats:p

Novel Aspects on the Regulation of Thylakoid Protein Phosphorylation

Thylakoid membrane proteins are phosphorylated by different enzymes, which are subject to different control mechanisms. Activation of the light harvesting complex (LHCII) kinase is signaled by the redox state of plastoquinone and the cytochrome b/f complex and modulated by the thiol reduction state. Phosphorylation of Photosystem II (PS II) proteins may involve kinase(s) associated with the PS II core complex that do not involve the cytochrome b/f complex. Exposure of the phosphoprotein phosphorylation site(s) to protein kinases is regulated by light-induced conformational changes. Thus, thylakoid protein phosphorylation is regulated at both the enzyme and substrate levels. Thylakoid protein dephosphorylation is also under regulatory control, involving interaction between an immunophilin and a membrane-bound phosphatase. The physiological significance of thylakoid protein phosphorylation is not fully understood. Phosphorylation of LHCII is suggested to have a dual role: i) regulation of the LHCII/PSII/PS I interaction, underlying the mechanism of energy transfer balance and ii) prevention of the light-induced aggregation of LHCII or LHCII-PS II complexes. The formation of such macrodomains may affect the dynamics of the thylakoid membrane, which requires unhindered lateral diffusion of integral protein complexes. Phosphorylation of PS II subunits appear to be essential for the repair of photodamage to its reaction center occurring during light stress conditions.</p

Marine Cyanobacteria Tune Energy Transfer Efficiency in their Light-harvesting Antennae by Modifying Pigment Coupling

AbstractPhotosynthetic organisms regulate energy transfer to fit to changes in environmental conditions. The biophysical principles underlying the flexibility and efficiency of energy transfer in the light-harvesting process are still not fully understood. Here we examine how energy transfer is regulatedin-vivo. We compare different acclimation states of the photosynthetic apparatus in a marine cyanobacterial species that is well adapted to vertical mixing of the ocean water column and identify a novel acclimation strategy for photosynthetic life under low light intensities. Antennae rods extend, as expected, increasing light absorption. Surprisingly, in contrast to what was known for plants and predicted by classic calculations, these longer rods transfer energy fasteri.e.more efficiently. The fluorescence lifetime and emission spectra dependence on temperature, at the range of 4-300K, suggests that energy transfer efficiency is tuned by modifying the energetic coupling strength between antennae pigments.</jats:p