Adaptation of biological membranes to temperature. The lack of homeoviscous adaptation in the sarcoplasmic reticulum

Temperature adaptation of biological membranes was examined by comparing the fragmented sarcoplasmic reticulum preparation of goldfish acclimated to different temperatures. Membrane fluidity was estimated using the fluorescence polarization technique. There was considerable variation between preparations, but no consistent differences in fluidity were observed between 5- and 25°C-acclimated goldfish, fish species adapted over an evolutionary period to arctic or desert temperatures, and rat. The fatty acid composition of the sarcoplasmic reticulum preparations of differently acclimated goldfish showed differences in the proportion of mono- and polyunsaturated fatty acids while the proportion of saturated fatty acids remained relatively constant. However, the fatty acid composition of sarcoplasmic reticulum phosphoglycerides became more unsaturated in the order rat, desert pupfish, arctic sculpin, which correlates with their respective environmental or body temperature. It is concluded that differences in membrane components other than fatty acids are important in determining membrane dynamic structure. The inability to demonstrate homeoviscous adaptation in sarcoplasmic reticulum is supported by other evidence suggesting that functions of the sarcoplasmic reticulum that are measured in vitro are not affected by such modifications of their phosphoglyceride fatty acid composition as occur during thermal acclimation.     

Iron and the antibotulinal efficacy of nitrite.

Combination of nitrite, isoascorbate, and ethylenediaminetetraacetic acid were compared for their antibotulinal efficacy in perishable canned cured meat. A dose response relationship of available iron to the antibotulinal efficacy of nitrite was demonstrated.     

Waiting with and without Recombination: The Time to Production of a Double Mutant

R. A. Fisher and H. J. Muller argued in the 1930s that a major evolutionary advantage of recombination is that it allows favorable mutations to be combined within an individual even when they first appear in different individuals. This effect is evaluated in a two-locus, two-allele model by calculating the average waiting time until a new genotypic combination first appears in a haploid population. Three approximations are developed and compared with Monte Carlo simulations of the Wright–Fisher process of random genetic drift in a finite population. First, a crude method, based on the deterministic accumulation of single mutants, produces a waiting time of 1/with no recombination and 1/with recombination between the two loci, whereμis the mutation rate,Nis the haploid population size, andRis the recombination rate. Second, the waiting time is calculated as the expected value of a heterogeneous geometric distribution obtained from a branching process approximation. This gives accurate estimates forNμlarge. The estimates for small values ofNμare considerably lower than the simulated values. Finally, diffusion analysis of the Wright–Fisher process provides accurate estimates forNμsmall, and the time scales of the diffusion process show a difference betweenR=0 and forR?0 of the same order of magnitude as seen in the deterministic analysis. In the absence of recombination, accurate approximations to the waiting time are obtained by using the branching process for highNμand the diffusion approximation for lowNμ. For lowNμthe waiting time is well approximated by 1/. WithR?0, the following dependence onNμis observed: ForNμ>1 the waiting time is virtually independent of recombination and is well described by the branching process approximation. ForNμ≈ the waiting time is well described by a simplified diffusion approximation that assumes symmetry in the frequencies of single mutants. ForNμ?1 the waiting time is well described by the diffusion approximation allowing asymmetry in the frequencies of single mutants. Recombination lowers the waiting time until a new genotypic combination first appears, but the effect is small compared to that of the mutation rate and population size. For largeNμ, recombination has a negligible effect, and its effect is strongest for smallNμ, in which case the waiting time approaches a fixed fraction of the waiting time forR=0. Free recombination lowers the waiting time to about 45% of the waiting time for absolute linkage for smallNμ. Selection has little effect on the importance of recombination in general.     

Involvement of PI 3-kinase and activated ERK in facilitating insulin-stimulated triacylglycerol synthesis in hepatocytes

Triacylglycerol synthesis was studied in hepatocytes isolated from fasted/refed rats by EDTA perfusion. Insulin induced a 1.5-fold increase in glucose incorporation into triacylglycerol. Insulin-stimulated triacylglycerol synthesis and insulin-stimulated protein kinase B/Akt activity were inhibited by the phosphatidylinositol 3-kinase inhibitors wortmannin and LY 294002, and the mitogen-activated protein kinase kinase inhibitor PD 98059. Inhibition of p70 ribosomal protein-S6 kinase with rapamycin was without effect. Insulin-stimulated pyruvate dehydrogenase activity was abolished by phosphatidylinositol 3-kinase inhibitors. No effect of insulin on acetyl CoA carboxylase activity was observed.     

Cloning and analysis of the mating type genes from Cochliobolus heterostrophus

Cochliobolus heterostrophus, a heterothallic Ascomycete, has a single mating type locus with two alternate forms called MAT-1 and MAT-2. MAT-1 was cloned by complementing a MAT-2 strain using a cosmid library from a MAT-1 strain and screening for a homothallic transformant. The cosmid recovered from this transformant was able to re-transform a MAT-2 strain to homothallism and MAT identity was proven by restriction fragment length polymorphism and conventional genetic mapping. All homothallic transformants could mate with either MAT-1 or MAT-2 strains, although the number of ascospores produced by self matings or crosses to MAT-2 strains was low. Progeny of selfed homothallic transformants were themselves homothallic. MAT-2 was cloned by probing a cosmid library from a MAT-2 strain with a fragment of insert DNA from the MAT-1 cosmid. A 1.5 kb subclone of either MAT-containing cosmid was sufficient to confer mating function in transformants. Examination of the DNA sequence of these subclones revealed that MAT-1 and MAT-2 contain 1297 by and 1171 bp, respectively, of completely dissimilar DNA flanked by DNA common to both mating types. Putative introns were found (one in each MAT gene) which, when spliced out, would yield open reading frames (ORFs) that occupied approximately 90% of the dissimilar DNA sequences. Translation of the MAT-1 ORF revealed similarity to the Neurospora crassa MATA, Podospora anserina mat?, and Saccharomyces cerevisiae MATα1 proteins; translation of the MAT-2 ORF revealed similarity to the N. crassa MATa, P. anserina mat+, and Schizosaccharomyces pombe mat-Mc proteins. These gene products are all proven or proposed DNA binding proteins. Those with similarity to MAT-2 are members of the high mobility group.     

Synaptic glutamate release by ventromedial hypothalamic neurons is part of the neurocircuitry that prevents hypoglycemia

The importance of neuropeptides in the hypothalamus has been experimentally established. Due to difficulties in assessing function in vivo, the roles of the fast-acting neurotransmitters glutamate and GABA are largely unknown. Synaptic vesicular transporters (VGLUTs for glutamate and VGAT for GABA) are required for vesicular uptake and, consequently, synaptic release of neurotransmitters. Ventromedial hypothalamic (VMH) neurons are predominantly glutamatergic and express VGLUT2. To evaluate the role of glutamate release from VMH neurons, we generated mice lacking VGLUT2 selectively in SF1 neurons (a major subset of VMH neurons). These mice have hypoglycemia during fasting secondary to impaired fasting-induced increases in the glucose-raising pancreatic hormone glucagon and impaired induction in liver of mRNAs encoding PGC-1alpha and the gluconeogenic enzymes PEPCK and G6Pase. Similarly, these mice have defective counterregulatory responses to insulin-induced hypoglycemia and 2-deoxyglucose (an antimetabolite). Thus, glutamate release from VMH neurons is an important component of the neurocircuitry that functions to prevent hypoglycemia.     

Specialized phloem parenchyma cells in Norway spruce (Pinaceae) bark are an important site of defense reactions

The bark anatomy of Norway spruce clones that were resistant or susceptible to Ceratocystis polonica, a bark-beetle-vectored fungal pathogen, was compared. The major difference concerned the axial parenchyma cells, called polyphenolic parenchyma (PP cells) because of their vacuolar deposits. The phenolic nature of the deposits was indicated by autofluorescence under blue light, and immunocytochemical studies demonstrating PP cells are enriched in phenylalanine ammonia lyase (EC 4.3.1.5), a key enzyme in phenolic synthesis. Susceptible-clone PP cells occurred as single rows filled with dense deposits. The resistant clone had 40% more PP cells, which occurred in rows two cells thick plus as individual cells scattered among the sieve cells and had lighter deposits. Trees inoculated with fungus were analyzed but a distinct fungal response could not be separated from the general wound response. In the resistant clone, phenolic bodies were reduced in size and density or disappeared completely 12 d after wounding, and PP cell size increased. The susceptible-clone phenolics and cell size changed only slightly. These data show that PP cells are active in synthesis, storage, and modification of phenolics in response to wounding, providing an important site of constitutive and inducible defenses.