DAX1 mutations map to putative structural domains in a deduced three-dimensional model.

The DAX1 protein is an orphan nuclear hormone receptor based on sequence similarity in the putative ligand-binding domain (LBD). DAX1 mutations result in X-linked adrenal hypoplasia congenita (AHC). Our objective was to identify DAX1 mutations in a series of families, to determine the types of mutations resulting in AHC and to locate single-amino-acid changes in a DAX1 structural model. The 14 new mutations identified among our 17 families with AHC brought the total number of families with AHC to 48 and the number of reported mutations to 42; 1 family showed gonadal mosaicism. These mutations included 23 frameshift, 12 nonsense, and six missense mutations and one single-codon deletion. We mapped the seven single-amino-acid changes to a homology model constructed by use of the three-dimensional crystal structures of the thyroid-hormone receptor and retinoid X receptor alpha. All single-amino-acid changes mapped to the C-terminal half of the DAX1 protein, in the conserved hydrophobic core of the putative LBD, and none affected residues expected to interact directly with a ligand. We conclude that most genetic alterations in DAX1 are frameshift or nonsense mutations and speculate that the codon deletion and missense mutations give insight into the structure and function of DAX1.     

Multiple trophic pathways support fish on floodplains of California's Central Valley

We used compound-specific isotope analysis of carbon isotopes in amino acids (AAs) to determine the biosynthetic source of AAs in fish from major tributaries to California's Sacramento-San Joaquin river delta (i.e., the Sacramento, Cosumnes and Mokelumne rivers). Using samples collected in winter and spring between 2016 and 2019, we confirmed that algae are a critical component of floodplain food webs in California's Central Valley. Results from bulk stable isotope analysis of carbon and nitrogen in producers and consumers were adequate to characterize a general trophic structure and identify potential upstream and downstream migration into our study site by American shad Alosa sapidissima and rainbow trout Oncorhynchus mykiss, respectively. However, owing to overlap and variability in source isotope compositions, our bulk data were unsuitable for conventional bulk isotope mixing models. Our results from compound-specific carbon isotope analysis of AAs clearly indicate that algae are important sources of organic matter to fish of conservation concern, such as Chinook salmon Oncorhynchus tshawytscha in California's Central Valley. However, algae were not the exclusive source of energy to metazoan food webs. We also revealed that other sources of AAs, such as bacteria, fungi and higher plants, contributed to fish as well. While consistent with the well-supported notion that algae are critical to aquatic food webs, our results highlight the possibility that detrital subsidies might intermittently support metazoan food webs.     

Complete nucleotide sequence of pSTK1, a cryptic plasmid from Bacillus stearothermophilus TK015

Summary The complete nucleotide sequence of pSTK1, a cryptic plasmid isolated from B. stearothermophilus TK015, has been determined. pSTK1 has been shown to be 1883 bp in length and contain three open reading frames (ORFs), one of which has a helix-turn-helix motif typical of DNA-binding proteins. Also identified was a region that can form an extensive secondary structure, which would show a high degree of similarity to palA, an origin for minus strand elongation in rolling circle replication.     

Identification of toxic metals in affected algal cells in assays of wastewaters

Chlorella fusca was the test species used in assays of two wastewaters, one characterized by a high concentration of free metal ions, and the other by high concentrations of free metal ions and chlorinated organic compounds. Fine structural damage in cells exposed for six days to the effluents was examined using an electron microscope. In the presence of organic toxicity, the affected cells displayed a characteristically large build-up of starch grains. At high metal concentrations, the chloroplasts were the only organelles altered. Within the chloroplast, there were disruptions of the thylakoidal membranes and enlargement of interthylakoidal spaces. Excess metals, detected by X-ray microanalysis in the polyphosphate (poly-P) bodies of cells where chloroplast damage was observed, were regarded as harmful. X-ray scanning of particulate material in effluent samples is proposed as a useful and low cost addition to surveillance programmes.     

The effect of exercise training on oxidative damage of lipids, proteins, and DNA in rat skeletal muscle: evidence for beneficial outcomes.

Moderate daily exercise is known to be beneficial to health, reducing risks of a number of age-related disorders. Molecular mechanisms that bring about these effects are not clear. In contrast, it has been claimed that some types of prolonged physical exertion are detrimental to health because active oxygen species are generated excessively by enhanced oxygen consumption. Using two age groups of rats, young (4 week) and middle aged (14 months), we investigated the effects of long-term swimming training on the oxidative status of phospholipids, proteins, and DNA. The concentration of thiobarbituric acid reactive substances and 4-hydroxynonenal protein adducts did not differ in the gastrocnemius muscle between exercised and nonexercised animals in the two age groups. The extent of carbonylation in a protein of molecular weight around 29 KDa and the amount of 8-hydroxydeoxyguanosine in nuclear DNA were smaller (p<.05) in the exercised rats than in the sedentary animals. Activities of DT-diaphorase (C1: 29.3+/-1.9; C2: 36.1+/-2.6; E1: 27.2+/-1.3; C2: 33.4+/-2.9 nmol/mg protein) and proteasome, a major proteolytic enzyme for oxidatively modified proteins were significantly higher in the exercised animals of both age groups (p<.05). The adaptive response against oxidative stress induced by moderate endurance exercise constitutes a beneficial effect of exercise.     

Catalytic control and coupling efficiency of the Escherichia coli FoF1 ATP synthase: influence of the Fo sector and epsilon subunit on the catalytic transition state

The rate-limiting transition state of steady-state ATP hydrolysis and synthesis reactions in the F(o)F(1) ATP synthase involves the rotation of the gamma, epsilon, and c subunits. To probe the role of the transport and coupling mechanisms in controlling catalysis, kinetic and thermodynamic parameters of ATP hydrolysis were determined for enzymes in the presence of the detergent lauryldimethylamine oxide (LDAO), which uncouples active transport and disables the inhibitory effect of the epsilon subunit. At 5 mM LDAO or greater, the inhibitory effects of epsilon subunit are abrogated in both purified F(1) and membranous F(o)F(1). In these conditions, LDAO solubilized F(o)F(1) has a higher k(cat) for ATP hydrolysis than F(1). These results indicate an influence of F(o) on F(1) even though catalysis is uncoupled from transport. The alpha(3)beta(3)gamma complex free of the epsilon subunit is activated at a lower concentration of 0.5 mM LDAO. Significantly, the gammaY205C mutant enzyme is similarly activated at 0.5 mM LDAO, suggesting that the mutant enzyme lacks epsilon inhibition. The gammaY205C F(o)F(1), which has a k(cat) for ATP hydrolysis 2-fold higher than wild type, has an ATP synthesis rate 3-fold lower than wild type, showing that coupling is inefficient. Arrhenius and isokinetic analyses indicate that enzymes that are free of epsilon subunit inhibition have a different transition-state structure from those under the influence of the epsilon subunit. We propose that the epsilon subunit is one of the factors that determines the proper transition-state structure, which is essential for efficient coupling.     

Phosphorylation of telokin by cyclic nucleotide kinases and the identification of in vivo phosphorylation sites in smooth muscle

The Ca(2+)-independent acceleration of dephosphorylation of the regulatory light chain of smooth muscle myosin and relaxation of smooth muscle by telokin are enhanced by cyclic nucleotide-activated protein kinase(s) [Wu et al. (1998) J. Biol. Chem. 273, 11362-113691. The purpose of this study was to determine the in vivo site(s) and in vitro rates of telokin phosphorylation and to evaluate the possible effects of sequential phosphorylation by different kinases. The in vivo site(s) of phosphorylation of telokin were determined in rabbit smooth muscles of longitudinal ileum and portal vein. Following stimulation of ileum with forskolin (20 microM) the serine at position 13 was the only amino acid to exhibit increased phosphorylation. Rabbit portal vein telokin was phosphorylated on both Ser-13 and -19 as a result of forskolin and GTPgammaS stimulation in vivo. Point mutation of Ser-13 (to Ala or Asp) abolished in vitro phosphorylation by cyclic nucleotide-dependent protein kinases.     

Association of insulin receptor substrate proteins with Bcl-2 and their effects on its phosphorylation and antiapoptotic function

Insulin receptor substrate (IRS) proteins are docking proteins that couple growth factor receptors to various effector molecules, including phosphoinositide-3 kinase, Grb-2, Syp, and Nck. Here we show that IRS-1 associates with the loop domain of Bcl-2 and synergistically up-regulates antiapoptotic function of Bcl-2. IRS-2 but not IRS-3 binds to Bcl-2, and IRS-1 associates with Bcl-XL but not with Bax or Bik. Overexpression of IRS-1 suppresses phosphorylation of Bcl-2 induced by stimulation with insulin, and the hypophosphorylation may lead to its enhanced antiapoptotic activity. The binding site for Bcl-2 is located on the carboxyl half-domain of IRS-1. IRS-3, which lacks the corresponding region, dominant-negatively abrogates the survival effects of IRS-1 and Bcl-2. For the antiapoptotic activity of IRS-1, binding to Bcl-2 is more critical than activating phosphoinositide-3 kinase. Our results indicate that IRS proteins transmit signals from the insulin receptor to Bcl-2, thus regulating cell survival probably through regulating phosphorylation of Bcl-2.     

Plume-tracking robots: a new application of chemical sensors

Many animals have the ability to search for odor sources by tracking their plumes. Some of the key features of this search behavior have been successfully transferred to robot platforms, although the capabilities of animals are still beyond the current level of sensor technologies. The examples described in this paper are (1) incorporating into a wheeled robot the upwind surges and casting used by moths in tracking pheromone plumes, (2) extracting useful information from the response patterns of a chemical sensor array patterned after the spatially distributed chemoreceptors of some animals, and (3) mimicking the fanning behavior of silkworm moths to enhance the reception of chemical signals by drawing molecules from one direction. The achievements so far and current efforts are reviewed to illustrate the steps to be taken toward future development of this technology.