Polyunsaturated fatty acid incorporation into plasmalogens in plasma membrane of glioma cells is preceded temporally by acylation in microsomes.

Plasmalogens (1-O-alk-1'-enyl-2-acyl-sn-glycero-3-phosphoethanolamine) are major phospholipids in many tissues and cells, particularly of neural origin. Using cultured C6 glioma cells and subcellular fractions isolated on Percoll gradients we investigated selectivity for esterification of several polyunsaturated fatty acids (PUFA) in the sn-2 position of plasmalogens compared to [1-14C]hexadecanol, representative of de novo synthesis of the ether-linked sn-1 position. In whole cells at a final concentration of 105 microM PUFA, 2-4 nmol plasmalogen/mg protein was labeled in 4 h and 10-14 nmol in 24 h, representing 8-15% and 35-50%, respectively, of initial plasmalogen mass. Incorporation of label from hexadecanol was lower than PUFA incorporation (20:5(n-3) greater than 20:4(n-6) greater than 18:3(n-3) much greater than 18:2(n-6)) suggesting deacylation-reacylation at the sn-2 position. Plasmalogens accounted for 50% of total cell ethanolamine phospholipids and 75% in plasma membrane. Using a novel, improved method for extraction of subcellular fractions containing Percoll, plasma membrane also was enriched in plasmalogen relative to microsomes (107.4 +/- 5.2 vs. 40.0 +/- 2.9 nmol/mg protein). Selectivity for esterification at the sn-2 position of plasmalogens with respect to chain length and unsaturation of the fatty acyl chain was similar in both subcellular fractions and reflected that of whole cells. Labeling of plasma membrane with PUFA and fatty alcohol lagged behind that of microsomes. Chase experiments in cells prelabeled with [1-14C]18:3(n-3) for 2 h showed no significant reduction of label in plasmalogen of any subcellular fraction although accumulation of label in the microsomal fraction was slowed initially. Reduction of plasmalogen label (40-50%) did occur in microsomes and plasma membrane when cells prelabeled for 24 h were switched to chase medium with or without chase fatty acid. Our data suggest that esterification of PUFA to plasmalogen may occur at the endoplasmic reticulum with subsequent translocation to plasma membrane resulting in accumulation of relatively stable pools of plasmalogen that are not readily accessible for deacylation-reacylation exchange with newly appearing PUFA. Alternatively, deacylation-reacylation may occur in a more stable phospholipid pool within the plasma membrane but would involve a slower process than at the endoplasmic reticulum.     

Genetic localization of Hao-1, blind-sterile (bs), and Emv-13 on mouse chromosome 2.

The blind-sterile (bs) mutation in the mouse was localized on Chromosome 2 between Hao-1 and Emv-13. N2 progeny from a backcross between congenic female 129.AKR-bs Emv-13 mice and (129.AKR-bs/bs x Mus musculus molossinus) F1 male mice were typed by analysis of isozyme variants for Hao-1, visible inspection for bs, and restriction fragment length polymorphism for Emv-13 and Emv-15. Comparison between markers on mouse Chromosome 2 and corresponding markers on human chromosomes suggest that the human homolog of bs will be located on 20q11-q13.     

Degumming of buel (Grewia optiva) bast fibres by pectinolytic enzyme from Bacillus sp. DT7

An alkalophilic and thermotolerant, pectinase-producing Bacillus sp. DT7 effectively removed pectic substances from buel (Grewia optiva) bast fibres. A novel combined (chemical and enzymatic) treatment was used to degum buel bast fibres, which was followed by the release of galacturonic acid (575 mol g^–1 dry fibres) and a decrease in dry weight (43%) of these fibres.     

Transitions from injection-drug-use-concentrated to self-sustaining heterosexual HIV epidemics: patterns in the international data

Background

Injecting drug use continues to be a primary driver of HIV epidemics in many parts of the world. Many people who inject drugs (PWID) are sexually active, so it is possible that high-seroprevalence HIV epidemics among PWID may initiate self-sustaining heterosexual transmission epidemics.

Methods

Fourteen countries that had experienced high seroprevalence (<20%) HIV epidemics among PWID and had reliable data for injection drug use (IDU) and heterosexual cases of HIV or AIDS were identified. Graphs of newly reported HIV or AIDS cases among PWID and heterosexuals were constructed to identify temporal relationships between the two types of epidemics. The year in which newly reported cases among heterosexuals surpassed newly reported cases among PWID, aspects of the epidemic curves, and epidemic case histories were analyzed to assess whether it was “plausible” or “highly unlikely” that the HIV epidemic among PWID might have initiated the heterosexual epidemic in each country.

Results

Transitions have occurred in 11 of the 14 countries. Two types of temporal relationships between IDU and heterosexual HIV epidemics were identified, rapid high incidence transitions vs. delayed, low incidence transitions. In six countries it appears “plausible” that the IDU epidemic initiated a heterosexual epidemic, and in five countries it appears “highly unlikely” that the IDU epidemic initiated a heterosexual epidemic. A rapid decline in incidence among PWID after the peak year of new cases and national income were the best predictors of the “highly unlikely” initiation of a heterosexual epidemic.

Discussion

Transitions from IDU concentrated epidemics to heterosexual epidemics are common in countries with high seroprevalence among PWID though there are distinct types of transitions. Interventions to immediately reduce HIV incidence among PWID may reduce the likelihood that an IDU epidemic may initiate a heterosexual epidemic.     

Effects of bacterial endotoxin on protecting copper-deficient rats from hyperoxia

The administration of very low doses of bacterial endotoxin protects rats during exposure to hyperoxia and is associated with the induction of lung antioxidant enzyme activities. Copper-deficient rats have increased susceptibility to O2 toxicity, which may be related to their decreased lung superoxide dismutase activity (SOD) or decreased plasma ceruloplasmin concentrations. To determine whether endotoxin can protect against hyperoxia in this susceptible model, we exposed copper-deficient and control rats to a fractional inspiratory concentration of O2 greater than 0.95 for 96 h after pretreatment with 500 micrograms/kg of bacterial endotoxin or phosphate-buffered saline (PBS). Mortality in the copper-deficient and control rats given PBS and exposed to O2 for 96 h was 100%. Copper-deficient rats died significantly earlier during the exposure than controls. No mortality occurred in either group treated with endotoxin and hyperoxia despite the decreased activity of copper-dependent enzymes in the copper-deficient rats. Copper-deficient rats treated with endotoxin and exposed to hyperoxia did increase lung Cu-Zn-SOD activity, but activity remained below levels found in air-exposed controls. Mn-SOD activity was found to be induced above air-exposed controls in the copper-deficient rats treated with endotoxin and exposed to hyperoxia. Hyperoxic exposure resulted in a marked increase in plasma ceruloplasmin concentrations in the control rats, but no increases in ceruloplasmin occurred in the copper-deficient animals. Endotoxin protects copper-deficient rats from hyperoxia despite their decreased lung Cu-Zn-SOD activity, and decreased plasma ceruloplasmin.     

The Anticancer Agent Di-2-pyridylketone 4,4-Dimethyl-3-thiosemicarbazone (Dp44mT) Overcomes Prosurvival Autophagy by Two Mechanisms: PERSISTENT INDUCTION OF AUTOPHAGOSOME SYNTHESIS AND IMPAIRMENT OF LYSOSOMAL INTEGRITY

Autophagy functions as a survival mechanism during cellular stress and contributes to resistance against anticancer agents. The selective antitumor and antimetastatic chelator di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone (Dp44mT) causes lysosomal membrane permeabilization and cell death. Considering the integral role of lysosomes in autophagy and cell death, it was important to assess the effect of Dp44mT on autophagy to further understand its mechanism of action. Notably, Dp44mT affected autophagy by two mechanisms. First, concurrent with its antiproliferative activity, Dp44mT increased the expression of the classical autophagic marker LC3-II as a result of induced autophagosome synthesis. Second, this effect was supplemented by a reduction in autophagosome degradation as shown by the accumulation of the autophagic substrate and receptor p62. Conversely, the classical iron chelator desferrioxamine induced autophagosome accumulation only by inhibiting autophagosome degradation. The formation of redox-active iron or copper Dp44mT complexes was critical for its dual effect on autophagy. The cytoprotective antioxidant N-acetylcysteine inhibited Dp44mT-induced autophagosome synthesis and p62 accumulation. Importantly, Dp44mT inhibited autophagosome degradation via lysosomal disruption. This effect prevented the fusion of lysosomes with autophagosomes to form autolysosomes, which is crucial for the completion of the autophagic process. The antiproliferative activity of Dp44mT was suppressed by Beclin1 and ATG5 silencing, indicating the role of persistent autophagosome synthesis in Dp44mT-induced cell death. These studies demonstrate that Dp44mT can overcome the prosurvival activity of autophagy in cancer cells by utilizing this process to potentiate cell death.     

A horizon scanning assessment of current and potential future threats to migratory shorebirds

We review the conservation issues facing migratory shorebird populations that breed in temperate regions and use wetlands in the non‐breeding season. Shorebirds are excellent model organisms for understanding ecological, behavioural and evolutionary processes and are often used as indicators of wetland health. A global team of experienced shorebird researchers identified 45 issues facing these shorebird populations, and divided them into three categories (natural, current anthropogenic and future issues). The natural issues included megatsunamis, volcanoes and regional climate changes, while current anthropogenic threats encompassed agricultural intensification, conversion of tidal flats and coastal wetlands by human infrastructure developments and eutrophication of coastal systems. Possible future threats to shorebirds include microplastics, new means of recreation and infectious diseases. We suggest that this review process be broadened to other taxa to aid the identification and ranking of current and future conservation actions.     

Methionine Adenosyltransferase and Ethionine Resistance in Saccharomyces cerevisiae

The methionine adenosyltransferase is repressed in Saccharomyces cerevisiae during growth in the presence of excess methionine. The relationship of this repression to the level of intracellular S-adenosylmethionine is discussed. In conjunction with these studies, an ethionine-resistant mutant has been investigated which has a low level of methionine adenosyltransferase under all conditions tested. The mechanism of ethionine resistance in the latter strain apparently depends on its inability to form large quantities of intracellular S-adenosylethionine. With respect to the methionine adenosyltransferase, there is no apparent interaction between ethionine-resistant and ethionine-sensitive alleles when both are present in the heterozygous diploid.     

Enhanced production of penicillin G acylase from a recombinant Escherichia coli

Penicillin G acylase (pac) gene was cloned into a stable asd ⁺ vector (pYA292) and expressed in Escherichia coli. This recombinant strain produced 1000 units penicillin G acylase g^–1 cell dry wt, which is 23-fold more than that produced by parental Escherichia coli ATCC11105. This enzyme was purified to 16 units mg^–1 protein by a novel two-step process.