Allosteric activation mechanism of the alpha1beta2gamma2 gamma-aminobutyric acid type A receptor revealed by mutation of the conserved M2 leucine

A conserved leucine residue in the midpoint of the second transmembrane domain (M2) of the ligand-activated ion channel family has been proposed to play an important role in receptor activation. In this study, we assessed the importance of this leucine in the activation of rat alpha1beta2gamma2 GABA receptors expressed in Xenopus laevis oocytes by site-directed mutagenesis and two-electrode voltage clamp. The hydrophobic conserved M2 leucines in alpha1(L263), beta2(L259), and gamma2(L274) subunits were mutated to the hydrophilic amino acid residue serine and coexpressed in all possible combinations with their wild-type and/or mutant counterparts. The mutation in any one subunit decreased the EC(50) and created spontaneous openings that were blocked by picrotoxin and, surprisingly, by the competitive antagonist bicuculline. The magnitudes of the shifts in GABA EC(50) and picrotoxin IC(50) as well as the degree of spontaneous openings were all correlated with the number of subunits carrying the leucine mutation. Simultaneous mutation of the GABA binding site (beta2Y157S; increased the EC(50)) and the conserved M2 leucine (beta2L259S; decreased the EC(50)) produced receptors with the predicted intermediate agonist sensitivity, indicating the two mutations affect binding and gating independently. The results are discussed in light of a proposed allosteric activation mechanism.     

Prevalence,comorbidities and mortality of toxic shock syndrome in children and adults in the USA

Toxic Shock Syndrome (TSS), a superantigen‐mediated illness, is characterized by rash, hypotension and multi‐organ dysfunction. Predictors of TSS and related morbidity and mortality are poorly defined. In this study, data on 61,959,084 hospitalizations from the 2003–2012 Nationwide Inpatient Sample, a 20% stratified sample of US hospitalizations, were analyzed and ICD‐9‐CM coding used to identify 4491 hospitalizations with a diagnosis of TSS. Incidence, in‐hospital mortality rate, comorbidities, length of stay and costs of care attributable to TSS were determined. In multivariate survey logistic regression models, TSS was associated with female sex (adjusted odds ratio [95% confidence interval], 1.54 [1.48–1.60]), younger age (0–17 years, 2.17 [2.06–2.29]; 40–59: 0.53 [0.50–0.56]; 60–79: 0.28 [0.26–0.30]; 80+: 0.13 [0.11–0.14] compared with 18–39) and race/ethnicity (black, 0.63 [0.59–0.67]; Hispanic: 0.60 [0.56–0.64]; Asian, 1.11 [1.00–1.11]; and other, 0.83 [0.75–0.92] compared with white). Patients with TSS had a three‐fold greater cost of care (mean: $36,656 ± 942) and length of stay (LOS) (mean: 10.65 ± 0.23 days) than patients without TSS. Shared predictors of increased LOS and costs in patients with TSS were male sex; age 40–79 years; Black, Hispanic, Asian and other race/ethnicity; and more than one chronic condition. Predictors of in‐hospital mortality included respiratory failure (13.66 [11.37–16.43]), liver disease/failure (3.36 [2.59–4.34]), chickenpox (91.26 [27.74–300.25]), coagulopathy (2.14 [1.85–2.48]), and higher age. In conclusion, there are significant racial/ethnic, socioeconomic, and comorbid disparities in the incidence and mortality of TSS in adults and children in the USA.

     

Biodecolourization of azo and triphenylmethane dyes by <Emphasis Type="Italic">Dichomitus squalens</Emphasis> and <Emphasis Type="Italic">Phlebia</Emphasis> spp

Nine white-rot fungal strains were screened for biodecolourization of brilliant green, cresol red, crystal violet, congo red and orange II. Dichomitus squalens, Phlebia fascicularia and P. floridensis decolourized all of the dyes on solid agar medium and possessed better decolourization ability than Phanerochaete chrysosporium when tested in nitrogen-limited broth medium. Journal of Industrial Microbiology & Biotechnology (2002) 28, 201–203 DOI: 10.1038/sj/jim/7000222 Received 12 July 2001/ Accepted in revised form 22 October 2001     

Phylogenetic evidence for horizontal transmission of group I introns in the nuclear ribosomal DNA of mushroom-forming fungi

Group I introns were discovered inserted at the same position in the nuclear small-subunit ribosomal DNA (nuc-ssu-rDNA) in several species of homobasidiomycetes (mushroom-forming fungi). Based on conserved intron sequences, a pair of intron-specific primers was designed for PCR amplification and sequencing of intron-containing rDNA repeats. Using the intron-specific primers together with flanking rDNA primers, a PCR assay was conducted to determine presence or absence of introns in 39 species of homobasidiomycetes. Introns were confined to the genera Panellus, Clavicorona, and Lentinellus. Phylogenetic analyses of nuc-ssu-rDNA and mitochondrial ssu-rDNA sequences suggest that Clavicorona and Lentinellus are closely related, but that Panellus is not closely related to these. The simplest explanation for the distribution of the introns is that they have been twice independently gained via horizontal transmission, once on the lineage leading to Panellus, and once on the lineage leading to Lentinellus and Clavicorona. BLAST searches using the introns from Panellus and Lentinellus as query sequences retrieved 16 other similar group I introns of nuc-ssu-rDNA and nuclear large-subunit rDNA (nuc-lsu-rDNA) from fungal and green algal hosts. Phylogenetic analyses of intron sequences suggest that the mushroom introns are monophyletic, and are nested within a clade that contains four other introns that insert at the same position as the mushroom introns, two from different groups of fungi and two from green algae. The distribution of host lineages and insertion sites among the introns suggests that horizontal and vertical transmission, homing, and transposition have been factors in intron evolution. As distinctive, heritable features of nuclear rDNAs in certain lineages, group I introns have promise as phylogenetic markers. Nevertheless, the possibility of horizontal transmission and homing also suggest that their use poses certain pitfalls.      

Effect of I,25-dihydroxycholecalciferol in renal osteodystrophy.

A 23-year-old man with medullary cystic disease had been undergoing hemodialysis for 5 years and had become confined to a wheelchair because of renal osteodystrophy. He was treated with 125-dihydroxycholecalciferol, 2.0 mug (later 1.0 mug) three times a week, administered by way of the venous end of the dialysis machine. Within 1 month bone pain lessened and his ability to stand and walk improved. By 3 months he was walking short distances and by 5 months, long distances. Calcium balance was near zero before treatment and was strongly positive during treatment. Bone mineral content in the lower femur, measured by photon absorptiometry, increased at a rate of 32.2% per year. In contrast, 26 other patients on long-term hemodialysis had a mean loss of bone mineral content of 14.0% per year. Radiographs taken during treatment showed a decrease in subperiosteal bone resorption and healing of a pseudofracture. A significant decrease in the mean serum alkaline phosphatase value was noted during treatment, but no significant changes in mean serum calcium or phosphorus values were seen.     

City-wide screening for urinary abnormalities in schoolgirls

Screening for urinary tract infection was carried out in 23,427 schoolgirls, aged 5 to 14 years, using Uricult and, for hematuria, glycosuria and proteinuria using Hema-combistix. Cultures of 10⁵ colonies per ml. or more on two occasions were obtained in 2.3% and a positive culture was confirmed by the family physician using standard culture techniques in 82.7% of cases, giving an overall incidence of infection of 1.9%. Fifty-eight percent of these children had no previous history of any urinary tract symptoms. Of the infected group 9.5% had pyelonephritic scarring, 58.7% chronic cystitis and 58.7% urethral stenosis. Two additional cases had unilateral ureteropelvic junction obstruction with hydronephrosis. Reflux occurred in 26.6% of those investigated by voiding cystogram. In 58% of cases the urinary tract infection was not accompanied by significant proteinuria, hematuria or pyuria.Proteinuria was detected on two occasions in 1.6% of the children and confirmed by the family physician in 33% of cases, giving an overall incidence of 0.5%. In this group 9.2% had evidence of pyelonephritic scarring without a positive urine culture.Hematuria was detected on two occasions in 0.6% of the children and was confirmed by the family physician in 53%, giving an overall incidence of 0.3%. Only one case with pyelonephritic scarring was seen in this group.Of the 25 cases with pyelonephritic changes only six had been previously diagnosed radiologically.Four previously unrecognized diabetics were also detected.     

Observations on the diagenetic behavior of arsenic in a deep coastal sediment

Vertical profiles of total dissolved arsenic, manganese and iron, pH, Eh and rates of sulfate reduction were determined in a freshly-collected box core from a 335m depth station in the Laurentian Trough. The relationships observed between the profiles were further examined in the laboratory by measuring these same parameters with time in surficial sediment slurries as the Eh decreased in response to biological activity or chemical alteration.Both field and laboratory observations have shown that arsenic is released predominantly as As(III) into reducing sediment porewaters. This occurs after the dissolution of manganese oxides and at the same time as the dissolution of iron oxyhydroxides and the onset of sulfate reduction. Laboratory experiments indicated that sulfate reduction and the production of sulfide ions are not solely responsible for the release of arsenic to the porewaters, although this process is necessary to create and maintain a highly reducing environment conducive to rapid iron dissolution.The diagenesis of arsenic in Laurentain Trough sediments involves the simultaneous release of arsenic and iron at a subsurface depth, followed by its removal from porewaters by precipitation and adsorption reactions after migration by diffusion along concentration gradients. A qualitative model is presented to describe the behavior of arsenic in coastal marine sediments.Present address: Department of Geological Sciences, McGill University, 3450 UniversityStreet, Montreal, Quebec H3A 2A7, Canada     

Rainfall-driven sex-ratio genes in African buffalo suggested by correlations between Y-chromosomal haplotype frequencies and foetal sex ratio

Background  

The Y-chromosomal diversity in the African buffalo (Syncerus caffer) population of Kruger National Park (KNP) is characterized by rainfall-driven haplotype frequency shifts between year cohorts. Stable Y-chromosomal polymorphism is difficult to reconcile with haplotype frequency variations without assuming frequency-dependent selection or specific interactions in the population dynamics of X- and Y-chromosomal genes, since otherwise the fittest haplotype would inevitably sweep to fixation. Stable Y-chromosomal polymorphism due one of these factors only seems possible when there are Y-chromosomal distorters of an equal sex ratio, which act by negatively affecting X-gametes, or Y-chromosomal suppressors of a female-biased sex ratio. These sex-ratio (SR) genes modify (suppress) gamete transmission in their own favour at a fitness cost, allowing for stable polymorphism.