The effect of cytochrome oxidase on lipid chain dynamics A nanosecond fluorescence depolarization study

Molecular motions in membranes composed of purified cytochrome oxidase (EC 1.9.3.1) and synthetic lipid (l-α-dimyristoylphosphatidylcholine or l-α-dioleoylphosphatidylcholine) at various ratios were investigated with a lipophilic fluorescent probe 1,6-diphenyl-1,3,5-hexatriene. Nanosecond fluorescence depolarization kinetics of the probe showed that the rod-shaped probe molecules perform a fast wobbling motion (restricted rotation) in all membranes studied, presumably reflecting the motion of lipid acyl chains. At temperatures where the pure lipid was in the liquid-crystalline phase, presence of cytochrome oxidase reduced the angular range of the wobbling motion, whereas its rate; the wobbling diffusion constant, was unaffected. On the other hand, incorporation of the protein into lipid in the gel phase resulted in the increase in the wobbling diffusion constant while the range of the wobbling motion remained the same. A time-dependent view of lipid dynamics that accounts for the above findings, as well as the results of recent electron spin resonance and nuclear spin resonance studies of protein-lipid interactions, is proposed.     

S4 protein Sll1252 is necessary for energy balancing in photosynthetic electron transport in Synechocystis sp. PCC 6803

Sll1252 was identified as a novel protein in photosystem II complexes from Synechocystis sp. PCC 6803. To investigate the function of Sll1252, the corresponding gene, sll1252, was deleted in Synechocystis 6803. Despite the homology of Sll1252 to YlmH, which functions in the cell division machinery in Streptococcus, the growth rate and cell morphology of the mutant were not affected in normal growth medium. Instead, it seems that cells lacking this polypeptide have increased sensitivity to Cl(-) depletion. The growth and oxygen evolving activity of the mutant cells was highly suppressed compared with those of wild-type cells when Cl(-) and/or Ca(2+) was depleted from the medium. Recovery of photosystem II from photoinhibition was suppressed in the mutant. Despite the defects in photosystem II, in the light, the acceptor side of photosystem II was more reduced and the donor side of photosystem I was more oxidized compared with wild-type cells, suggesting that functional impairments were also present in cytochrome b(6)/f complexes. The amounts of cytochrome c(550) and cytochrome f were smaller in the mutant in the Ca(2+)- and Cl(-)-depleted medium. Furthermore, the amount of IsiA protein was increased in the mutant, especially in the Cl(-)-depleted medium, indicating that the mutant cells perceive environmental stress to be greater than it is. The amount of accompanying cytochrome c(550) in purified photosystem II complexes was also smaller in the mutant. Overall, the Sll1252 protein appears to be closely related to redox sensing of the plastoquinone pool to balance the photosynthetic electron flow and the ability to cope with global environmental stresses.     

Identification and origin of the germline stem cells as revealed by the expression of nanos-related gene in planarians

The planarian's remarkable regenerative ability is thought to be supported by the stem cells (neoblasts) found throughout its body. Here we report the identification of a subpopulation of neoblasts, which was revealed by the expression of the nanos-related gene of the planarian Dugesia japonica, termed Djnos. Djnos-expressing cells in the asexual planarian were distributed to the prospective ovary or testes forming region in the sexual planarian. During sexualization, Djnos-expressing cells produce germ cells, suggesting that in the asexual state these cells were kept as germline stem cells for the oogonia and spermatogonia. Interestingly, the germline stem cells were indistinguishable from the neoblasts by morphology and X-ray sensitivity and did not seem to contribute to the regeneration at all. Germline stem cells initially appear in the growing infant planarian, suggesting that germline stem cells are separated from somatic stem cells in the planarian. Thus, planarian neoblasts can be classified into two groups; somatic stem cells for regeneration and tissue renewal, and germline stem cells for production of germ cells during sexualization. However, Djnos-positive cells appeared in the newly formed trunk region from the head piece, suggesting that somatic stem cells can convert to germline stem cells.     

Influence of <Emphasis Type="Italic">Flammulina velutipes</Emphasis> mycelia culture conditions on antimicrobial metabolite production

Enokipodins A, B, C, and D are α-cuparene-type sesquiterpenoids antimicrobial metabolites produced in the stationary stage of Flammulina velutipes mycelia development in malt extract broth. This study assessed the influence of nutritional and environmental factors on F. velutipes mycelia culture for the production of these metabolites. The mycelia growth and antimicrobial activity were assessed by determining dry matter and the diffusion in agar method, respectively. The best F. velutipes mycelia growth was observed in dextrose potato broth, and greater antimicrobial metabolite production occurred in complete Pontecorvo’s culture medium. Environmental modifications, such as a rise in temperature from 25° to 37°C on the 15th day of F. velutipes mycelia culture in malt extract and peptone broth, also optimized antimicrobial metabolite production. The metabolites produced in these treatments were correlated with the enokipodins A and B in thin-layer chromatography (TLC) and the antifungal activity test by TLC bioautography. This study showed that there was no correlation between biomass production and antimicrobial metabolite production, but there may be a correlation between culture medium composition and enokipodins biosynthesis.     

Zellweger syndrome and a microdeletion of the proximal long arm of chromosome 7

Summary A 16-day-old girl with Zellweger syndrome and a chromosomal rearrangement, 46,XX,del(7)(q11.22q11.23), is reported. The diagnosis was confirmed by marked deficiencies of peroxisomal -oxidation enzymes and dihydroxyacetone phosphate acyltransferase activities in rectal cells and fibroblasts obtained by biopsy and in hepatic cells obtained at autopsy. This is the first report of Zellweger syndrome associated with a chromosomal arrangement, a microdeletion of chromosome 7. A tentative gene assignment to 7q11 is suggested.     

Changes in membrane surface properties of hepatic peroxisomes of rats under several conditions as determined by partition in aqueous polymer two-phase systems

Changes in membrane surface properties of hepatic peroxisomes of rats under several conditions were observed by aqueous polymer two-phase systems, which contained 6% (w/w) dextran T 500, 6% (w/w) polyethyleneglycol 4000, 250 mmol sucrose/kg and various concentrations of sodium phosphate buffer. The partition of peroxisomes into the upper phase depended to a large extent on their membrane surface charge. The cross-points of peroxisomes shifted from 5.55 to 5.25 and 5.2 after the administration of clofibrate and aspirin for 2 weeks, respectively, although that of alloxan-diabetic rat peroxisomes was not altered. The hydrophobic properties of peroxisomes, examined by means of a partition containing polyethyleneglycol monostearate, were altered by diabetes and starvation, but no change occurred in rats treated with clofibrate or aspirin. In the liver of rats fed a high-fat diet, the partition of peroxisomes was the same as that of the control. These findings indicate that hypolipidemic drugs such as clofibrate and aspirin induce the proliferation of peroxisomes and lead to the alteration of the surface charge of peroxisomal membranes. Diabetes or fasting lead to an alteration mainly of the hydrophobic properties. Both changes are probably due to alteration of content and/or composition of the proteins and the phospholipids in peroxisomal membrane under the conditions used.     

Mucopolysaccharidosis IVA: Identification of a Common Missense Mutation I113F in the N-Acetylgalactosamine-6-Sulfate Sulfatase Gene

Mucopolysaccharidosis IVA is an autosomal recessive lysosomal storage disorder caused by a deficiency of N-acetylgalactosamine-6-sulfate sulfatase. The recent isolation and characterization of cDNA and genomic sequences encoding GALNS has facilitated identification of the molecular lesions that cause MPS IVA. We identified a common missense mutation among Caucasian MPS IVA patients. The mutation was originally detected by SSCP, and successive sequencing revealed an A→T transversion at nt 393. This substitution altered the isoleucine at position 113 to phenylalanine (I113F) in the 622 amino acid GALNS protein and was associated with a severe phenotype in a homozygote. Compound heterozygotes with one I113F-allele mutation have a wide range of clinical phenotypes. Transfection experiments in GALNS-deficient fibroblasts revealed that the mutation drastically reduces the enzyme activity of GALNS. Allele-specific oligonucleotide or SSCP analysis indicated that this mutation accounted for 22.5% (9/40) of unrelated MPS IVA chromosomes from 23 Caucasian patients, including 6 consanguineous cases. Of interest, the I1e 113→Phe substitution occurred in only Caucasian MPS IVA patients and in none of the GALNS alleles of 20 Japanese patients. These findings identify a frequent missense mutation among MPS IVA patients of Caucasian ancestry, that results in severe MPS IVA when homoallelic, and will facilitate molecular diagnosis of most such patients and identification of heterozygous carriers. In addition to this common mutation, 10 different point mutations and 2 small deletions were detected, suggesting allelic heterogeneity in GALNS gene.     

cDNA cloning and partial sequencing of homeobox genes in Dugesia japonica

In the freshwater planarian Dugesia japonica, four types of cDNAs of homeobox-containing genes have been isolated by screening a cDNA library using a homeobox guessmer. Partial sequencing analysis of two types of cDNAs revealed that one was a homolog of Dth2 which is a homeobox gene in Dugesia tigrina and another was similar to Distal-less gene in Drosophila. This suggests that planarians have many homeobox genes.