The importance of arginine mutation for the evolutionary structure and function of phenylalanine hydroxylase gene

Phenylalanine hydroxylase (PAH) gene mutations were investigated in 23 (46 alleles) unrelated phenylketonuria (PKU) patients in Cukurova region. First, all exons of PAH gene were screened by denaturing high performance liquid chromatography (DHPLC), and then, the suspicious samples were analyzed by direct sequencing technique. Consequently, the following results were obtained: IVS10-11g-->a splicing mutation in 27/46 (58.7%), R261Q mutation in 7/46 (15.2%) and E178G, R243X, R243Q, P281L, Y386C, R408W mutations, each found in the frequency of 2/46 (4.3%). In many countries, Arginine mutations have the highest frequency among PAH gene mutations in PKU patients. Although, CpG dinucleotids are effective in mutations resulting in arginine changes, this finding originated from the studies on the causes of mutations rather than the studies on the importance of arginine amino acid. In our analyses, we have detected that a majority of mutations causing a change in arginine and other amino acids concentrated in exon 7 comprising the catalytic domain (residues 143-410) of PAH gene. Several studies has emphasized the role of arginine amino acid; with the following outcomes; arginine repetition is significant for RNA binding proteins, and for histon proteins in eukaryotic gene expression, and also arginine repetition occurring in the structure of signal recognition particle's (SRPs) as a consequence of post-translational processes is very important in terms of gene expression. Therefore, the role of arginine amino acid in PAH gene is rather remarkable in that it shows the role of amino acids in the protein/RNA interaction that has started in the evolutionary process and is still preserved and maintained in the motif formation of active domain structure due to its strong binding properties. Thus, such properties imply that both arginine amino acid and exon 7 is of great significance with regards to the structure and function of the PheOH enzyme.     

RNA modifications by oxidation: A novel disease mechanism?

The past decade has provided exciting insights into a novel class of central (small) RNA molecules intimately involved in gene regulation. Only a small percentage of our DNA is translated into proteins by mRNA, yet 80% or more of the DNA is transcribed into RNA, and this RNA has been found to encompass various classes of novel regulatory RNAs, including, e.g., microRNAs. It is well known that DNA is constantly oxidized and repaired by complex genome maintenance mechanisms. Analogously, RNA also undergoes significant oxidation, and there are now convincing data suggesting that oxidation, and the consequent loss of integrity of RNA, is a mechanism for disease development. Oxidized RNA is found in a large variety of diseases, and interest has been especially devoted to degenerative brain diseases such as Alzheimer disease, in which up to 50-70% of specific mRNA molecules are reported oxidized, whereas other RNA molecules show virtually no oxidation. The iron-storage disease hemochromatosis exhibits the most prominent general increase in RNA oxidation ever observed. Oxidation of RNA primarily leads to strand breaks and to oxidative base modifications. Oxidized mRNA is recognized by the ribosomes, but the oxidation results in ribosomal stalling and dysfunction, followed by decreased levels of functional protein as well as the production of truncated proteins that do not undergo proper folding and may result in protein aggregation within the cell. Ribosomal dysfunction may also signal apoptosis by p53-independent pathways. There are very few reports on interventions that reduce RNA oxidation, one interesting observation being a reduction in RNA oxidation by ingestion of raw olive oil. High urinary excretion of 8-oxo-guanosine, a biomarker for RNA oxidation, is highly predictive of death in newly diagnosed type 2 diabetics; this demonstrates the clinical relevance of RNA oxidation. Taken collectively the available data suggest that RNA oxidation is a contributing factor in several diseases such as diabetes, hemochromatosis, heart failure, and β-cell destruction. The mechanism involves free iron and hydrogen peroxide from mitochondrial dysfunction that together lead to RNA oxidation that in turn gives rise to truncated proteins that may cause aggregation. Thus RNA oxidation may well be an important novel contributing mechanism for several diseases.     

Implications of body condition on the unsustainable predation rates of endangered mountain caribou

Both top-down and bottom-up processes influence herbivore populations, and identifying dominant limiting factors is essential for applying effective conservation actions. Mountain caribou are an endangered ecotype of woodland caribou (Rangifer tarandus caribou) that have been declining, and unsustainable predation has been identified as the proximate cause. To investigate the role of poor nutrition, we examined the influence of sex, season, age class, and available suitable habitat (i.e., old-growth forest>140 years) per caribou on bone marrow fat content of caribou that died (n = 79). Sex was the only strong predictor of marrow fat. Males that died during and post rut had lower marrow fat than females or males at other times of year. Old-growth abundance per caribou, season, and age class did not predict marrow fat. Caribou killed by predators did not have less marrow fat than those that died in accidents, suggesting that nutritionally stressed caribou were not foraging in less secure habitats or that predators selected nutritionally stressed individuals. Marrow fat in endangered and declining populations of mountain caribou was similar to caribou in other, more viable populations. Our results support previous research suggesting that observed population declines of mountain caribou are due to excessive predation that is not linked to body condition.     

Fine mapping of QTLs for rice grain yield under drought reveals sub-QTLs conferring a response to variable drought severities

Fine-mapping studies on four QTLs, qDTY(2.1), qDTY(2.2), qDTY(9.1) and qDTY(12.1), for grain yield (GY) under drought were conducted using four different backcross-derived populations screened in 16 experiments from 2006 to 2010. Composite and bayesian interval mapping analyses resolved the originally identified qDTY(2.1) region of 42.3 cM into a segment of 1.6 cM, the qDTY(2.2) region of 31.0 cM into a segment of 6.7 cM, the qDTY(9.1) region of 32.1 cM into two segments of 9.4 and 2.4 cM and the qDTY(12.1) region of 10.6 cM into two segments of 3.1 and 0.4 cM. Two of the four QTLs (qDTY(9.1) and qDTY(12.1)) having effects under varying degrees of stress severity showed the presence of more than one region within the original QTL. The study found the presence of a donor allele at RM262 within qDTY(2.1) and RM24334 within qDTY(9.1) showing a negative effect on GY under drought, indicating the necessity of precise fine mapping of QTL regions before using them in marker-assisted selection (MAS). However, the presence of sub-QTLs together in close vicinity to each other provides a unique opportunity to breeders to introgress such regions together as a unit into high-yielding drought-susceptible varieties through MAS.     

11beta-hydroxysteroid dehydrogenase 2 activity is elevated in severe obesity and negatively associated with insulin sensitivity

Alterations in glucocorticoid (GC) metabolism may contribute to the development of obesity and insulin resistance. We aimed to study the role of 11beta-hydroxysteroid dehydrogenase type 2 (11beta-HSD2) in human adiposity, paying special attention to the association between altered GC metabolism and insulin sensitivity. In 24-h urine samples of 72 extremely obese (mean BMI 45.5 +/- 1.1 kg/m(2)), but otherwise healthy patients urinary free cortisol (UFF), urinary free cortisone (UFE), tetrahydrocortisol (THF), 5alpha-tetrahydrocortisol (5alpha-THF), and tetrahydrocortisone (THE) were quantified by radioimmunoassay. The sum of the three major tetrahydrometabolites is an estimate for daily GC secretion, and the sum of UFF and UFE represents potentially bioactive-free-GCs. Thirty healthy lean subjects (BMI 22.3 +/- 0.3 kg/m(2)) served as controls. In obese subjects, absolute daily GC secretion and the potentially bioactive-free-GCs were significantly (P < 0.005) higher than in lean controls (11.8 +/- 0.7 vs. 8.0 +/- 0.6 mg/d; and 171.8 +/- 11.2 vs. 117.6 +/- 9.2 mug/d, respectively). However, when these values were corrected for body surface area (BSA), significant differences were no longer detectable. While enzyme activity indices for 5alpha-reductase and 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) were similar in lean and obese subjects, 11beta-HSD2 was markedly elevated in adiposity (3.7 +/- 0.2 vs. 2.1 +/- 0.1; P < 0.0001). This increase was accompanied by a significant reduction in UFF excretion corrected for BSA (16.5 +/- 1.2 vs. 21.7 +/- 2.0 mug/d/m(2); P = 0.0222). Besides, 11beta-HSD2 activity was significantly correlated with insulin sensitivity (P = 0.0262). When body size is accounted for, both adrenal GC secretion and potentially bioactive-free-GCs are indistinguishable between lean and extremely obese subjects. However in obesity, the kidney appears to intensify its supply of the direct substrate cortisone for extrarenal 11beta-HSD1, which may fuel visceral adiposity and insulin resistance.     

Synthesis of rhamnogalacturonan I fragments by a modular design principle

The improved syntheses of methyl 2-O-acetyl-3-O-benzyl-alpha-L-rhamnopyranoside (12) and 1,2-di-O-acetyl-3-O-benzyl-alpha-L-rhamnopyranose (15), which were used as glycosyl acceptor and donor, respectively, are described. Glycosylation of the O-4 position of both rhamnose derivatives with 2,3,4,6-tetra-O-benzoyl-alpha-D-galactopyranosyl bromide (26) provided disaccharides 27 and 29. After partial deprotection of 27 and coupling of the resulting 28 with disaccharide 19, tetrasaccharide 31 was obtained. Furthermore, transforming of 29 into the corresponding bromide 30 and coupling with galacturonates 16 and 32 provided trisaccharides 33 and 34, respectively, which could be regarded as building blocks of ramified rhamnogalacturonan fragments. The preparation of tetra- (21) and hexasaccharide (25) of rhamnogalacturonan I is reported to demonstrate the feasibility of the synthesis of larger pectin fragments using the modular design principle with this type of building blocks.     

The AAE14 gene encodes the Arabidopsis o-succinylbenzoyl-CoA ligase that is essential for phylloquinone synthesis and photosystem-I function

Phylloquinone is the one-electron carrier at the A₁ site of photosystem I, and is essential for photosynthesis. Arabidopsis mutants deficient in early steps of phylloquinone synthesis do not become autotrophic and are seedling lethals, even when grown on sucrose-supplemented media. Here, we identify acyl-activating enzyme 14 (AAE14, At1g30520) as the o -succinylbenzoyl-coenzyme A (OSB-CoA) ligase acting in phylloquinone synthesis. Three aae14 mutant alleles, identified by reverse genetics, were found to be seedling lethal, to contain no detectable phylloquinone (< 0.1 pmol mg⁻¹ fresh weight) compared with 10 pmol mg⁻¹ fresh weight in wild-type leaves, and to accumulate OSB. AAE14 was able to restore menaquinone biosynthesis when expressed in an Escherichia coli mutant disrupted in the menE gene that encodes the bacterial OSB-CoA ligase. Weak expression of an AAE14 transgene in mutant plants (controlled by the uninduced XVE promoter) resulted in chlorotic, slow-growing plants that accumulated an average of 4.7 pmol mg⁻¹ fresh weight of phylloquinone. Inducing the XVE promoter in these plants, or expressing an AAE14 transgene under the control of the CaMV 35S promoter, led to full complementation of the mutant phenotype. aae14 -mutant plants were also able to synthesize phylloquinone when provided with 1,4-dihydroxy-2-naphthoate, an intermediate in phylloquinone synthesis downstream of the OSB-CoA ligase reaction. Expression of an AAE14:GFP reporter construct indicated that the protein accumulated in discrete foci within the chloroplasts. This and other evidence suggests that the enzymes of phylloquinone synthesis from isochorismate may form a complex in the chloroplast stroma to facilitate the efficient channeling of intermediates through the pathway.