The Effects of Aluminum on the Influx of Calcium,Potassium, Ammonium,Nitrate, and Phosphate in an Aluminum-Sensitive Cultivar of Barley (Hordeum vulgare L.)

The mechanism by which aluminum interferes with ion influx is not known. In this study, the effects of aluminum on the influx of the cations calcium, potassium, and ammonium and the anions nitrate and phosphate were measured in an aluminum-sensitive cultivar of barley (Hordeum vulgare L.). Aluminum (100 [mu]M) was found to inhibit the influx of the cations calcium (69%), ammonium (40%), and potassium (13%) and enhancing the influx of the anions nitrate (44%) and phosphate (17%). Aluminum interfered with the binding of the cations in the cell wall by the same order of magnitude as their respective influxes, whereas phosphate binding was strongly enhanced. The results are consistent with a mechanism whereby aluminum binds to the plasma membrane phospholipids, forming a positively charged layer that influences ion movement to the binding sites of the transport proteins. A positive charge layer would retard the movement of cations and increase the movement of anions to the plasma membrane in proportion to the charges carried by these ions.     

Characterization of the non-specific lipid transfer protein EP2 from carrot (Daucus carota L.)

The extracellular protein EP2 was previously identified as non-specific lipid transfer protein based on its cDNA-derived amino acid sequence. Here, the purification of the EP2 protein from the medium of somatic embryo cultures is described. After two cycles of ion-exchange and gel permeation chromatography, a single silver-stained protein band with an apparent molecular mass of 10 kDa was observed on SDS-PAGE. This protein band was recognized by the antiserum raised against a EP2--galactosidase fusion-protein. Employing a fluorescent phospholipid analog, it was shown that the purified EP2 protein is capable of binding phospholipids and is able to enhance their transfer between artificial membranes. Employing a gel permeation assay, it could be demonstrated that the EP2 protein is also capable of binding palmitic and oleic acid as well as oleyl-CoA. Because in plants these fatty acids are used as precursor molecules for cutin, these results are in support of the proposed role of the EP2 protein to transport cutin monomers from their site of synthesis through the cell wall of epidermal cells to sites of cutin polymerization.     

In vitro oxidative inactivation of glutathione S-transferase from a freeze tolerant reptile

We have previously reported that when garter snakesThamnophis sirtalis parietalis, a freeze tolerant species, were exposed to 5 h freezing at –2.5° C organs showed increases in the activities of anti-oxidant enzymes, especially catalase in skeletal muscle. This was interpreted to be an adaptation to deal with the potentially injurious postischemic situation of thawing. The present work analyzesin vitro oxidative inactivation of a possible target of postischemic-induced free radical damage, the secondary anti-oxidant defense glutathione-S transferase, and the protective role of endogenous catalase. Approximately 50% of GST activity from snake muscle homogenates was lost within 2 min after addition of H₂O₂ plus Fe(II) (0.4–2 mM) in media containing azide whereas addition of iron alone resulted in no damaging effects. The opposing effects of dimethyl sulfoxide and EDTA in modifying this process strongly suggested the involvement of ·OH radicals in the GST inactivation. A partial recovery of the activity was promoted by mercaptoethanol, indicating that sulphydryl groups oxidation participate in the mechanism of GST inactivation. Pre-incubation of the reaction media containing H₂O₂ caused protection of the GST activity only in the absence of azide, indicating that endogenous catalase modulates the extent of oxyradical damage. The protective pre-incubation effect was more efficacious when employing homogenates from lung and liver, organs that have higher catalase activities, as well as homogenates from freezing-exposed muscle (that show an 80% increase in catalase activity, compared with control). The protection against GST inactivation observed in muscle from frozen snakes demonstrates that increased anti-oxidant defenses during freezing exposure can be a key factor in controllingin vitro oxyradical damage. The implications for natural freeze tolerance are discussed.     

High prevalence of a point mutation in the porphobilinogen deaminase gene in Dutch patients with acute intermittent porphyria

Acute intermittent porphyria (AIP) is an autosomal dominant disease characterized by a deficiency of porphobilinogen deaminase (PBGD). Up to now 14 different mutations have been described. In an effort to investigate the molecular epidemiology of AIP we have undertaken a systematic study of different exons of the PBGD gene from a large number of unrelated patients. Here, exon 8 from 82 unrelated Dutch and French AIP patients was examined using single strand confirmation polymorphism analysis (SSCP) after polymerase chain reaction (PCR) amplification. A single base mutation, C to T, at position 346 of the sequence coding for PBGD was observed in 15 Dutch families but in only 1 French family. A simple PCR assay is described to facilitate the diagnosis of this common mutation at the DNA level.     

Paternal duplication of chromosome 5q11.2–5q14 in a male born with craniostenosis,ear tags,kidney dysplasia and several other anomalies

A de novo duplication of the proximal part of the long arms of chromosome 5 was found in a male born with craniostenosis, ear tags and kidney dysplasia. The nature of the chromosomal aberration was defined by fluorescence in situ hybridization and the orgin of the duplication was traced by polymorphic DNA markers. A comparison is made with the published cases showing similar duplications in the long arm of chromosome 5.