Multi-platform metabolomics analyses of a broad collection of fragrant and non-fragrant rice varieties reveals the high complexity of grain quality characteristics.

The quality of rice in terms not only of its nutritional value but also in terms of its aroma and flavour is becoming increasingly important in modern rice breeding where global targets are focused on both yield stability and grain quality. In the present paper we have exploited advanced, multi-platform metabolomics approaches to determine the biochemical differences in 31 rice varieties from a diverse range of genetic backgrounds and origin. All were grown under the specific local conditions for which they have been bred and all aspects of varietal identification and sample purity have been guaranteed by local experts from each country. Metabolomics analyses using 6 platforms have revealed the extent of biochemical differences (and similarities) between the chosen rice genotypes. Comparison of fragrant rice varieties showed a difference in the metabolic profiles of jasmine and basmati varieties. However with no consistent separation of the germplasm class. Storage of grains had a significant effect on the metabolome of both basmati and jasmine rice varieties but changes were different for the two rice types. This shows how metabolic changes may help prove a causal relationship with developing good quality in basmati rice or incurring quality loss in jasmine rice in aged grains. Such metabolomics approaches are leading to hypotheses on the potential links between grain quality attributes, biochemical composition and genotype in the context of breeding for improvement. With this knowledge we shall establish a stronger, evidence-based foundation upon which to build targeted strategies to support breeders in their quest for improved rice varieties.

Natural selection results in conservation of HIV-1 integrase activity despite sequence variability.

BACKGROUND: Integration of the HIV genome by integrase is absolutely required for productive infection. OBJECTIVE: To determine the role of natural selection on HIV integrase biology. DESIGN: To study the activities of HIV integrases from a limited panel of North American clinical isolates from HIV-infected patients and to compare these proteins with integrases from two laboratory adapted reference strains (HI(VIIIRF) and HIV(NL4--3)). METHODS: HIV was isolated and the particle-associated RNA was reverse transcribed and sequenced. Replication kinetics of molecularly cloned viruses containing each variant integrase were studied in tissue culture. The mutant integrase proteins were expressed, purified and specific activities of the enzymes were derived for both 3' end-processing and disintegration reactions. RESULTS: Despite 3--5% variability in integrase at the amino acid level, viruses showed no statistically significant differences in growth kinetics compared with the reference HIV(NL4--3) virus and only minor differences were observed in 3' end-processing and disintegration activities. All integrase proteins demonstrated similar sensitivity to an integrase inhibitor l-chicoric acid. CONCLUSIONS: These results demonstrate that integrase genes derived from HIV-infected individuals can differ from reference sequences but these mutations do not result in loss of function, including susceptibility to an integrase inhibitor; therefore, integrase remains an attractive target for antiviral drug design, as mutability appears to be restricted by function.

The sidekick gene, a member of the immunoglobulin superfamily, is required for pattern formation in the Drosophila eye.

In the Drosophila eye imaginal disc the photoreceptor cells (R cells) differentiate according to a precise spatial and temporal order. The sidekick (sdk) gene is necessary to prevent extra R cells from differentiating during eye disc development. The extra cell appears between R3 and R4 early in R cell clusters and is most likely the result of the mystery cell inappropriately differentiating as an R cell. Mosaic analysis shows that sdk is required neither in the R cells nor in the extra cell, suggesting that sdk is necessary in the surrounding undifferentiated cells. The sdk gene codes for a protein that is a member of the immunoglobulin superfamily, having six immunoglobulin domains, thirteen fibronectin repeats and a transmembrane domain. The protein structure is consistent with its participation in cell-cell interaction during eye development.

orthodenticle is required for photoreceptor cell development in the Drosophila eye.

Drosophila photoreceptor cells (R cells) develop from the eye imaginal during the third instar larval stage and acquire their adult morphology during pupation. We show that orthodenticle (otd), a homeobox gene, is required for R-cell morphogenesis during pupation. otdUV-insensitive (otduvi) is a hypomorphic allele of otd that only affects R-cell development. The R-cell rhabdomeres are disorganized in otduvi, and there is a disruption of proximal-distal development in the eye. The otd genomic structure was determined and resulted in the identification of a deletion in the third intron of otduvi. Sequences encompassing this deletion are able to direct expression of the lacZ reporter gene at all stages of the developing visual system, including the photosensitive cells of Bolwig's organ, the ocelli, and the adult eye. The third intron enhancer is the primary regulatory element controlling otd in the R cells and is not under the control of the glass gene.

Shark, a Src homology 2, ankyrin repeat, tyrosine kinase, is expressed on the apical surfaces of ectodermal epithelia.

Tyrosine kinases, ankyrin repeats, and Src homology 2 domains play central roles in developmental processes. The cloning of a cDNA for Shark, a single protein that possesses all three domains, is described. During Drosophila embryogenesis, Shark is expressed exclusively by ectodermally derived epithelia and is localized preferentially to the apical surface of these cells. This apical localization persists, even as tissues undergo complex invaginations, moving from the external surface of embryos to form internal structures, but expression is lost when cells lose their polarity. This pattern closely mimics the expression of Crumbs, a protein necessary for proper organization of ectodermal epithelia. Shark's structure and localization pattern suggest that it functions in a signaling pathway for epithelial cell polarity, possibly transducing the Crumbs signal.

Mercury burden of human fetal and infant tissues.

UNLABELLED: The total mercury concentrations in the liver (Hg-L), the kidney cortex (Hg-K) and the cerebral cortex (Hg-C) of 108 children aged 1 day-5 years, and the Hg-K and Hg-L of 46 fetuses were determined. As far as possible, the mothers were interviewed and their dental status was recorded. The results were compared to mercury concentrations in the tissues of adults from the same geographical area. The Hg-K (n = 38) and Hg-L (n = 40) of fetuses and Hg-K (n = 35) and Hg-C (n = 35) of older infants (11-50 weeks of life) correlated significantly with the number of dental amalgam fillings of the mother. The toxicological relevance of the unexpected high Hg-K of older infants from mothers with higher numbers of dental amalgam fillings is discussed. CONCLUSION: Future discussion on the pros and cons of dental amalgam should not be limited to adults or children with their own amalgam fillings, but also include fetal exposure. The unrestricted application of amalgam for dental restorations in women before and during the child-bearing age should be reconsidered.

Cell-cell interaction in the Drosophila retina: the bride of sevenless gene is required in photoreceptor cell R8 for R7 cell development.

Genetic mosaic studies indicate that environmental cues play a critical role in photoreceptor cell (R-cell) development in the Drosophila compound eye. Recent analysis of the sevenless gene suggests that its product, a cell surface protein containing a putative intracellular domain homologous to tryosine kinases, is a receptor for a signal specifying an R7-specific pathway of cellular development. We report the identification of a second genetic locus, bride of sevenless (boss) required for the development of R7. Genetic mosaic analysis demonstrates that boss expression is required in the R8 cell for a neighboring cell to become R7. However, the development of R1-R6 and R8 is boss-independent. These data indicate a central role for R8 in directing ommatidial assembly and are consistent with boss encoding the ligand recognized by the sevenless receptor.

Analysis of mutants in chaoptin, a photoreceptor cell-specific glycoprotein in Drosophila, reveals its role in cellular morphogenesis.

Monoclonal antibody 24B10 (MAb24B10) specifically stains photoreceptor neurons in D. melanogaster. It recognizes a 160 kd glycoprotein localized to the extracellular face of the plasma membrane. Using an immunoscreen, we identified two mutations in the encoding gene that cause microvillar disorganization in developing rhabdomeres and disruption of the closely apposed membranes of adjacent cells. In accordance with the mutant phenotype, we have renamed this genetic locus chaoptic and the encoded glycoprotein, chaoptin. Immunoelectron microscopy indicates that chaoptin is distributed along the length of the microvillus. This localization and the morphological abnormalities in mutants support the hypothesis that chaoptin may mediate adhesion between closely apposed membranes. In principle, the immunoscreen utilized here can be used to identify mutations in any gene in Drosophila for which antibodies to the gene product are available.

Chaoptin, a cell surface glycoprotein required for Drosophila photoreceptor cell morphogenesis, contains a repeat motif found in yeast and human.

Chaoptin is a photoreceptor cell-specific membrane protein. Analysis of chaoptin mutants demonstrates that this glycoprotein plays a critical role in photoreceptor cell morphogenesis. We have deduced chaoptin's primary structure from the cDNA sequence and examined its membrane topology. Chaoptin is largely composed of 41 potentially amphipathic repeats. Remarkably homologous repeats have been reported in both yeast and human, suggesting their general importance as a structural and/or functional motif. Chaoptin synthesized in vitro is associated with rough endoplasmic reticulum microsomal membrane in an integral fashion, consistent with its extraction characteristics in vivo. The resistance to proteolytic digestion of in vitro synthesized and processed chaoptin suggests that it is primarily localized to the extracellular leaflet of the lipid bilayer. These data are consistent with the proposal that chaoptin is involved in the adhesion between photoreceptor cell membranes.

Glucocorticoid-regulated expression from the 5'-flanking region of the rat alpha 1-acid glycoprotein gene. Requirement for ongoing protein synthesis.

The induction of alpha 1-acid glycoprotein (AGP) mRNA by glucocorticoids in rat hepatoma cells requires ongoing protein synthesis. Here we show that the 5'-flanking region of the AGP gene confers glucocorticoid responsiveness on the expression of heterologous coding sequences. Moreover, the induction of beta-globin mRNA directed by the AGP promoter is inhibited by concurrent treatment with cycloheximide, thereby suggesting that the requirement for protein synthesis is mediated through 5'-flanking sequences. Analysis of the effects of varying durations of cycloheximide treatment indicates that both the endogenous AGP gene and the transfected AGP-beta-globin chimeric gene are induced normally by dexamethasone during the first 1-2h of concurrent treatment. In addition, pretreatment with cycloheximide yields a decrease in the subsequent induction of both beta-globin and AGP RNAs. These data support the notion that a pre-existing and labile protein, perhaps in concert with the glucocorticoid receptor, acts through the 5'-flanking region of the AGP gene to stimulate transcription from the AGP promoter.

Acute phase mediators and glucocorticoids elevate alpha 1-acid glycoprotein gene transcription.

Acute phase mediators and glucocorticoids increase the synthesis of the acute phase protein alpha 1-acid glycoprotein, also known as orosomucoid, by inducing the hepatic level of its mRNA. Concurrently the acute phase response depresses the hepatic synthesis of albumin and alpha 2u-globulin and their mRNA levels. Present transcriptional studies in isolated liver nuclei demonstrate that turpentine-induced acute phase mediators simultaneously enhance transcription of the alpha 1-acid glycoprotein gene and diminish transcription of albumin and alpha 2u-globulin genes; parallel alterations in the hepatic level of the corresponding mRNAs ensue. The present transcriptional studies also demonstrate that administration of dexamethasone to adrenalectomized rats dramatically elevates the rate of transcription of the alpha 1-acid glycoprotein gene as well as the alpha 2u-globulin and the albumin genes, leading to elevations in alpha 1-acid glycoprotein and alpha 2u-globulin hepatic mRNA levels. Thus, hepatic alpha 1-acid glycoprotein mRNA levels are predominantly regulated in vivo at the transcriptional level by glucocorticoids as well as by acute phase mediators.

Rat alpha 1-acid glycoprotein. Gene sequence and regulation by glucocorticoids in transfected L-cells.

We have cloned and sequenced the rat gene coding for the acute phase reactant protein alpha 1-acid glycoprotein in order to determine which sequences are necessary for its regulation by glucocorticoids and which sequences are responsible for the sensitivity of this regulation to protein synthesis inhibitors. The gene contains six exons, as determined from the alpha 1-acid glycoprotein cDNA sequence, and five introns. Primer extension and S1 nuclease experiments have shown that there are two transcriptional start sites 4 base pairs apart. After cotransfection into mouse L-cells, the gene retains its inducibility by glucocorticoids, indicating that the sequences required for induction are within or around the gene. The induction of alpha 1-acid glycoprotein levels in transfected L-cells is sensitive to protein synthesis inhibitors, implying that in addition to the glucocorticoid receptor another protein(s) is necessary for full induction.

Left atrial dimension on the frontal thoracic radiograph: a method for assessing left atrial enlargement.

Left atrium size on the frontal radiograph was assessed by measuring the distance from the middle of the "double density" to the left main bronchus in 148 normal patients and 48 consecutive patients with echocardiographic evidence of left atrial enlargement. The measurement was less than 7.0 cm in 96% of normal patients and greater than 7.0 cm in 90% of patients with left atrial enlargement. This measurement is more easily obtainable than measurements and signs requiring a cardiac series, and is usually readily available for sequential evaluations of left atrial size. It is a less reliable gauge of left atrial size in children.

Patent ductus arteriosus in preterm infants with idiopathic respiratory distress syndrome. Radiographic and echocardiographic evaluation.

Clinical, radiographic, echocardiographic and operative findings were evaluated in 55 preterm infants with idiopathic respiratory distress syndrome (IRDS) complicated by patent ductus arteriosus (PDA). Pulmonary plethora was detected prior to age seven days in 52 infants, and prior to murmur detection in 42 infants. In those with large shunts, only 35% had cardiomegaly while 78% had a significant increase in cardiothoracic ratio (C/T) on sequential radiographs. Moreover, within 48 hours after ligation, 91% of infants had a significant decrease in C/T. Echocardiographic left atrial to aortic ratio (LA/Ao) was elevated in 71% with large shunts. In one patient with a large shunt there was neither a sequential increase in C/T nor an increased LA/Ao. Severity of left-to-right shunting across a PDA in the newborn was reliably gauged by combined radiographic and echocardiographic evaluation. Either modality alone failed to reflect the presence of a large volume shunt in a number of infants.

Salicylate-induced pulmonary edema.

A case of pulmonary edema secondary to salicylate intoxication is described. The pulmonary wedge pressure was normal, excluding cardiogenic pulmonary edema. Thus salicylate intoxication should be considered as a rare cause in the differential diagnosis of pulmonary edema with a normal heart size. The pulmonary edema resolved gradually over 8 days. Literature relative to salicylate-induced pulmonary edema is reviewed.

Bilateral pulmonary hilar lymphadenopathy. An unusual manifestation of metastatic renal cell carcinoma.

Four patients with bilateral pulmonary hilar adenopathy secondary to lymphangitic spread from renal cell carcinoma were examined. Two additional cases had adenopathy secondary to nasopharyngeal carcinoma. Patients may initially present with bilateral pulmonary lymphadenopathy or as late as 3 1/2 years after the discovery of the primary renal tumor. The mechanism of lymphangitic spread probably is related to reflux of tumor emboli from the thoracic duct into the bronchomediastinal trunks because of incompetent lymphatic valves. In one case gallium imaging demonstrated bilateral hilar isotopic uptake as well as periaortic uptake.

Perforation of the nasopharynx by nasogastric intubation: a rare cause of left pleural effusion and pneumomediastinum.

A case of nasogastric tube perforation of the posterior nasopharynx producing a left pleural effusion and pneumomediastinum is reported. Findings which led to the correct diagnosis were the rapid appearance of a left pleural effusion simultaneous with the initiation of tube feedings, the intrathoracic location of the nasogastric tube, and demonstration of perforation of the posterior nasopharynx. Potentially hazardous intubation techniques are discussed.