Cytosolic thioredoxin reductase 1 is required for correct disulfide formation in the ER

Folding of proteins entering the secretory pathway in mammalian cells frequently requires the insertion of disulfide bonds. Disulfide insertion can result in covalent linkages found in the native structure as well as those that are not, so‐called non‐native disulfides. The pathways for disulfide formation are well characterized, but our understanding of how non‐native disulfides are reduced so that the correct or native disulfides can form is poor. Here, we use a novel assay to demonstrate that the reduction in non‐native disulfides requires NADPH as the ultimate electron donor, and a robust cytosolic thioredoxin system, driven by thioredoxin reductase 1 (TrxR1 or TXNRD1). Inhibition of this reductive pathway prevents the correct folding and secretion of proteins that are known to form non‐native disulfides during their folding. Hence, we have shown for the first time that mammalian cells have a pathway for transferring reducing equivalents from the cytosol to the ER, which is required to ensure correct disulfide formation in proteins entering the secretory pathway.     

Detection of two loci involved in (1-->3)-beta-glucan (curdlan) biosynthesis by Agrobacterium sp. ATCC31749, and comparative sequence analysis of the putative curdlan synthase gene

Genes essential for the production of a linear, bacterial (1-->3)-beta- glucan, curdlan, have been cloned for the first time from Agrobacterium sp. ATCC31749. The genes occurred in two, nonoverlapping, genomic fragments that complemented different sets of curdlan( crd )-deficient transposon-insertion mutations. These were detected as colonies that failed to stain with aniline blue, a (1-->3)-beta-glucan specific dye. One fragment carried a biosynthetic gene cluster (locus I) containing the putative curdlan synthase gene, crdS, and at least two other crd genes. The second fragment may contain only a single crd gene (locus II). Determination of the DNA sequence adjacent to several locus I mutations revealed homology to known sequences only in the cases of crdS mutations. Complete sequencing of the 1623 bp crdS gene revealed highest similarities between the predicted CrdS protein (540 amino acids) and glycosyl transferases with repetitive action patterns. These include bacterial cellulose synthases (and their homologs), which form (1-->4)-beta-glucans. No similarity was detected with putative (1-->3)- beta-glucan synthases from yeasts and filamentous fungi. Whatever the determinants of the linkage specificity of these beta-glucan synthases might be, these results raise the possibility that (1-->3)-beta-glucans and (1-->4)-beta-glucans are formed by related catalytic polypeptides.      

Modeling expression quantitative trait loci in data combining ethnic populations

Background  

Combining data from different ethnic populations in a study can increase efficacy of methods designed to identify expression quantitative trait loci (eQTL) compared to analyzing each population independently. In such studies, however, the genetic diversity of minor allele frequencies among populations has rarely been taken into account. Due to the fact that allele frequency diversity and population-level expression differences are present in populations, a consensus regarding the optimal statistical approach for analysis of eQTL in data combining different populations remains inconclusive.     

Emission analysis of RE3+ (RE = Sm,Dy):B2O3‐TeO2‐Li2O‐AlF3 glasses

This article reports on the optical properties of 0.5% mol of Sm³⁺, Dy³⁺ ion‐doped B₂O₃‐TeO₂‐Li₂O‐AlF₃ (LiAlFBT) glasses. The glass samples were characterized by optical absorption and emission spectra. Judd‐Ofelt theory was applied to analyze the optical absorption spectra and calculate the intensity parameters and radiative properties of the emission transitions. The emission spectra of Sm³⁺ and Dy³⁺:LiAlFBT glasses showed a bright reddish‐orange emission at 598 nm (⁴G_5/2 → ⁶H_7/2) and an intense yellow emission at 574 nm (⁴F_9/2 → ⁶H_13/2), respectively. Full width at half maximum (FWHM), stimulated emission cross section, gain bandwidth and optical gain values were also calculated to extend the applications of the Sm³⁺ and Dy³⁺:LiAlFBT glasses. Copyright © 2012 John Wiley & Sons, Ltd.     

Rates of nuclear and cytoplasmic mitochondrial DNA sequence divergence in mammals

Differential rates of nucleotide substitution among different gene segments and between distinct evolutionary lineages is well documented among mitochondrial genes and is likely a consequence of locus-specific selective constraints that delimit mutational divergence over evolutionary time. We compared sequence variation of 18 homologous loci (15 coding genes and 3 parts of the control region) among 10 mammalian mitochondrial DNA genomes which allowed us to describe different mitochondrial evolutionary patterns and to produce an estimation of the relative order of gene divergence. The relative rates of divergence of mitochondrial DNA genes in the family Felidae were estimated by comparing their divergence from homologous counterpart genes included in nuclear mitochondrial DNA (Numt, pronounced "new might"), a genomic fossil that represents an ancient transfer of 7.9 kb of mitochondrial DNA to the nuclear genome of an ancestral species of the domestic cat (Felis catus). Phylogenetic analyses of mitochondrial (mtDNA) sequences with multiple outgroup species were conducted to date the ancestral node common to the Numt and the cytoplasmic (Cymt) mtDNA genes and to calibrate the rate of sequence divergence of mitochondrial genes relative to nuclear homologous counterparts. By setting the fastest substitution rate as strictly mutational, an empirical "selective retardation index" is computed to quantify the sum of all constraints, selective and otherwise, that limit sequence divergence of mitochondrial gene sequences over time.      

Slow oscillations as a probe of the dynamics of the locus coeruleus-frontal cortex interaction in anesthetized rats

Multiunit or single unit activity recorded simultaneously from frontal cortex (FC) and locus coeruleus (LC) under ketamine anesthesia revealed that both regions show slow oscillatory activity, together or separately. If, however, both regions are engaged in this oscillatory activity, there is a systematic relationship between their phases with peak LC firing always following FC firing by 200–400 ms. This was confirmed by cross-correlational analyses, which indicated that the two structures temporarily form a resonant system. The FC-LC resonant state is, however, loose enough to remain open to other intrinsic or extrinsic influences, keeping the measured frequencies of oscillations at each site slightly different, as demonstrated by a delailed analysis of the autocorrelograms. An injection of lidocaine at the frontal cortex site, while sharply reducing the prefrontal activity to essentially zero, leads to an increase of the LC activity and to a modification of the shape of the LC autocorrelogram, but does not change appreciably the phase relationship between the activity in the two structures during the diminishing activity in FC.     

Liver scarring induced by polychlorinated biphenyl administration to mice previously treated with diethylnitrosamine

Exposure of mice for 8 weeks to drinking water containing diethylnitrosamine (DEN) was accompanied by alterations in hepatocyte structure and varying degrees of liver nonparenchymal cell (NPC) proliferation. Eighteen and a half weeks after cessation of DEN exposure, there was a 47% incidence of hepatocellular nodules. Centrilobular hepatocyte hypertrophy occurred consistently in mice given intraperitoneal injections of the polychlorinated biphenyl (PCB) mixture Aroclor 1254. PCB administration to mice previously treated with DEN was not accompanied by increases in gross liver nodule incidence above that induced by DEN, but many more developing microscopic nodules within the liver were observed in DEN-treated mice given Aroclor 1254 than in mice treated only with DEN. Aroclor 1254 administration over a 16-week period to mice previously treated with DEN was accompanied by an 83% incidence of severe distortion of liver structure resulting from nodule formation, uneven patterns of hepatocyte growth, and extensive deposition of scar tissue containing proliferating bile ducts. Morphological evidence of intestinal metaplasia was observed in proliferating bile duct-like structures during an early stage of liver adenofibrosis.     

Multiple actins in drosophila melanogaster

The tissue and developmental specificities of the three Drosophila isoactins, originally identified in primary myogenic cultures and in the permanent Schneider L-2 cell line, have been investigated. Of these three isoactins (I, II, and III), actins I and II are stable and actin III is unstable. Two-dimensional polyacrylamide gel electrophoretic analyses of total cellular extracts after 1-h [(35)S]methionine pulses were performed on a large variety of embryonic, larval, and adult muscle and nonmuscle tissues. The results suggest that isoactins II and III are generalized cellular actins found in all drosophila cell types. Actin I, on the other hand, is muscle-associated and is found exclusively in supercontractile muscle (such as larval body wall and larval and adult viscera) including primary myogenic cell cultures. Although actin I synthesis is not detectable during very early embryogenesis, it is detectable by 25 h and actin I is a major stable actin in all larval muscle tissues. Actin I is synthesized in reduced amounts relative to the other actins in late third instar larvae but is again a major product of actin synthesis in the adult abdomen. A stable actin species with the same pI as actin III has been identified in the adult thorax and appears to be unique to flight muscle tissue. This new stable form of thoracic actin may be the result of a stabilization of the actin III found in other tissues or may be an entirely separate gene product.     

Selective inhibition of prostaglandin biosynthesis by gold salts and phenylbutazone

Gold salts and phenylbutazone selectively inhibit the synthesis of PGF_2α and PGE₂ respectively. Lowered production of one prostaglandin species is accompanied by an increased production of the other. Selective inhibition by these drugs was observed in the presence of adrenaline, reduced glutathione and copper sulphate under conditions when most anti-inflammatory compounds inhibited PGE₂ and PGF_2α syntheses equally. It is postulated that selective inhibitors may have a different mode of action in vivo and beneficial effects may be related to the endogenous ratio of PGE to PGF required for normal function.