Characterization of the highly active polyhydroxyalkanoate synthase of Chromobacterium sp. strain USM2

The synthesis of bacterial polyhydroxyalkanoates (PHA) is very much dependent on the expression and activity of a key enzyme, PHA synthase (PhaC). Many efforts are being pursued to enhance the activity and broaden the substrate specificity of PhaC. Here, we report the identification of a highly active wild-type PhaC belonging to the recently isolated Chromobacterium sp. USM2 (PhaC(Cs)). PhaC(Cs) showed the ability to utilize 3-hydroxybutyrate (3HB), 3-hydroxyvalerate (3HV), and 3-hydroxyhexanoate (3HHx) monomers in PHA biosynthesis. An in vitro assay of recombinant PhaC(Cs) expressed in Escherichia coli showed that its polymerization of 3-hydroxybutyryl-coenzyme A activity was nearly 8-fold higher (2,462 ± 80 U/g) than that of the synthase from the model strain C. necator (307 ± 24 U/g). Specific activity using a Strep2-tagged, purified PhaC(Cs) was 238 ± 98 U/mg, almost 5-fold higher than findings of previous studies using purified PhaC from C. necator. Efficient poly(3-hydroxybutyrate) [P(3HB)] accumulation in Escherichia coli expressing PhaC(Cs) of up to 76 ± 2 weight percent was observed within 24 h of cultivation. To date, this is the highest activity reported for a purified PHA synthase. PhaC(Cs) is a naturally occurring, highly active PHA synthase with superior polymerizing ability.     

Summer distribution of Red-legged Partridges Alectoris rufa in relation to water availability on Mediterranean farmland

The suggestion that the summer distribution of Red-legged Partridges Alectoris rufa in the Mediterranean region is determined by the availability of surface water was examined on the agricultural farm, Alto Alentejo, southern Portugal. Partridge coveys were surveyed between 15 July and 15 August in 1993 and 1994. Using a vector-based Geographic Information System, we assessed, for each covey location and for the locations of a double number of random points, the distance to the nearest water point, distance to field boundaries, distance to water lines and land use classes. Univariate comparisons were made between the two groups of locations, and three multivariate logistic models were fitted through forward stepwise selection to the 1993, 1994 and pooled data sets to estimate the probability of sighting partridge coveys in the study area. Distance to water was significantly lower for partridge locations than for random points in both years and was the only variable selected for all logistic models. Apart from water availability, Red-legged Partridge locations were also affected by land use and distance to field boundaries.     

MAP, a protein interacting with a tumor suppressor, merlin, through the run domain

Merlin (or schwannomin) is a tumor suppressor encoded by the neurofibromatosis type 2 gene. Many studies have suggested that merlin is involved in the regulation of cell growth and proliferation through interactions with various cellular proteins. To better understand the function of merlin, we tried to identify the proteins that bind to merlin using the yeast two-hybrid screening. Characterization of the positive clones revealed a protein of 749 amino acids named merlin-associated protein (MAP), which showed wide tissue distribution in Northern blot analysis. Sequence analysis revealed that MAP is a potential homologue of a yeast check-point protein, BUB2, and contains TBC, SH3, and RUN domains, thereby implicating its role in the Ras-like GTPase signal pathways. MAP and merlin were directly associated in vitro and in vivo, and colocalized in NIH3T3 cells. The RUN domain of MAP and the C-terminus of merlin appeared to be responsible for their interaction. MAP decreased the AP-1-dependent promoter activity additively with merlin in NIH3T3 cells. In addition, merlin and MAP synergistically reduced the colony formation of NIH3T3 cells. These results suggest that MAP may play a cooperative role in the merlin-mediated growth suppression of cells.     

Zea ribosomal repeat evolution and substitution patterns

Zea and Tripsacum nuclear ribosomal internal transcribed spacer (ITS) sequences were used to evaluate patterns of concerted evolution, rates of substitutions, patterns of methylation-induced deamination, and structural constraints of the ITS. ITS pseudogenes were identified by their phylogenetic position, differences in nucleotide composition, extensive deamination at ancestral methylation sites, and substitutions resulting in low-stability secondary RNA structures. Selection was important in shaping the kinds of polymorphisms and substitutions observed in the ITS. ITS substitution rates were significantly different among the Zea taxa. Deamination of cytosines at methylation sites was a potent mutation source, but selection appeared to maintain high methylation site density throughout the ribosomal repeat except for the gene promoter. Nucleotide divergence statistics identified selectively constrained regions at the 5' ends of the ITS1 and ITS2.      

Development and validation of microsatellite markers for the tiny dragonfly, <Emphasis Type="Italic">Nannophya pygmaea</Emphasis> (Odonata: Libellulidae), which is endangered in South Korea

The tiny dragonfly, Nannophya pygmaea (Odonata: Libellulidae), is listed as a second-degree endangered wild animal in South Korea. The application of molecular markers to assess genetic diversity and population relationships can provide information necessary to establish an effective conservation strategy. In this study, we developed 12 microsatellite markers specific to N. pygmaea using the NextSeq 500 platform. Forty individuals of N. pygmaea collected from three currently known localities in South Korea were genotyped to validate these markers and to preliminarily assess population genetic characteristics. The observed number of alleles, observed heterozygosity, and expected heterozygosity at a locus ranged from 2 to 9, 0.421–1.0, and 0.508–0.766 in a population with the largest sample size (20 individuals), respectively, thereby validating the suitability of the markers for population analysis. Five of 12 loci showed significant deviation from the Hardy–Weinberg equilibrium in the population. Our preliminary data indicate an absence of inbreeding in all populations and an absence of obvious genetic difference. The microsatellite markers developed in this study will be useful for studying the population genetics of N. pygmaea collected from other regions worldwide, including additional sites in South Korea.     

Concerted transpositions of mobile genetic elements coupled with fitness changes in Drosophila melanogaster

In an inbred low-activity (LA) strain of Drosophila melanogaster with a low level of fitness and a complex of inadaptive characters, in situ hybridization reveals an invariant pattern of distribution of three copia-like elements (mdg-1, mdg-3, and copia). Rare, spontaneous, multiple transpositions of mobile elements in the LA strain were shown to be coupled with a drastic increase of fitness. A changed pattern of various types of mobile elements was also observed on selecting the LA strain for higher fitness. High-fitness strains show transpositions of mobile elements to definite chromosomal sites ("hot spots"). Concerted changes in the location of three different mobile elements were found to be coupled with an increase of fitness. The mdg-1 distribution patterns were also examined in two low-fitness strains independently selected from the high-fitness ones. Fitness decrease was accompanied by mdg-1 excision from the hot spots of their location usually detected in the high-fitness strains. The results suggest the existence of a system of adaptive transpositions of mobile elements that takes part in fitness control.      

Socioeconomic Disparities in the Prevalence of Blepharoptosis in the South Korean Adult Population Based on a Nationwide Cross-Sectional Study

Purpose

We investigated the association between socioeconomic status (SES) and the prevalence of blepharoptosis in a representative South Korean population.

Methods

This cross-sectional study was based on data obtained in the Korea National Health and Nutrition Examination Survey from 2010 to 2012. In total, 17,178 Korean adults (7,261 men and 9,917 women) aged 19 years or older were enrolled. Blepharoptosis was defined as a marginal reflex distance 1 (MDR 1) lower than 2 mm. Household income and education level were used as indicators of SES. Univariate and multiple logistic regression analyses were conducted to analyze the relationship between SES and the prevalence of blepharoptosis.

Results

Household income was inversely associated with the prevalence of blepharoptosis in women [adjusted odds ratio (aOR) and corresponding 95% confidence interval (95% CI) was 1.894 (1.336, 2.685)], and educational level was inversely associated with blepharoptosis in both men and women [aORs and 95% CIs were 1.572 (1.113, 2.219) and 1.973 (1.153, 3.376), respectively]. After adjusting for household income and educational level, low SES was associated with a high prevalence of blepharoptosis in women only.

Conclusions

Socioeconomic disparities in the prevalence of blepharoptosis were found among women. Indeed, future research using a prospective design to determine the causal relationship between SES and blepharoptosis may identify SES as a risk factor for this condition.     

Comparison of Trifecta and Pentafecta Outcomes between T1a and T1b Renal Masses following Robot-Assisted Partial Nephrectomy (RAPN) with Minimum One Year Follow Up: Can RAPN for T1b Renal Masses Be Feasible?

Purpose/Objectives

To investigate the feasibility of RAPN on T1b renal mass by assessment of Trifecta and Pentafecta rate between T1a and T1b renal mass.

Materials/Methods

We retrospectively reviewed the medical records of 277 cases of RPN performed from 2006 to 2015. Sixty patients with clinically T1b renal masses (> 4cm and ≤ 7 cm) were identified, and from 180 patients with clinically T1a renal mass, 60 patients were matched with T1b renal mass by propensity score. Tumor complexity was investigated according to R.E.N.A.L nephrometry score. “Pentafecta” was defined as achievement of Trifecta (negative surgical margin, no postoperative complications and warm ischemia time of ≤ 25 minutes) with addition of over 90% estimated GFR preservation and no chronic kidney disease stage upgrading at 1 year postoperative period. Propensity score matching was performed by OneToManyMTCH. Logistic regression models were used to identify the variables which predict the Trifecta, and Pentafecta ac.

Results

Preoperative variables (age, sex, body mass index, ASA score) were similar between T1a and T1b after propensity score matching. The median R.E.N.A.L. nephrometry score was 8 vs 9 for T1a and T1b respectively (p<0.001). The median warm ischemia time was 20.1 min vs 26.2 min (p<0.001). Positive surgical margin rate was 5% vs 6.6% (p = 0.729) and overall complication rate of 13.3%. vs 15% (p = 0.793). The rate of achievement of Trifecta rate were 65.3% vs 43.3% (p = 0.017) and Pentafecta rate were 38.3% vs 26.7% (p = 0.172). For achievement of Pentafecta, R.E.N.A.L nephrometry score (HR 0.80; 95% CI (0.67–0.97); p = 0.031) was significant predictor of achieving Pentafecta. Subanalyis to assess the component of R.E.N.A.L nephrometry score, L component (location relative to the polar lines, HR 0.63; 95% CI (0.38–1.03); P = 0.064) was relatively important component for Pentafecta achievement.

Conclusions

The rate of Pentafecta after RAPN was comparable between T1a and T1b renal masses. RAPN is a feasible modality with excellent long term outcome for patients with larger renal mass (cT1b).