Absence of PTPN11 mutations in 28 cases of cardiofaciocutaneous (CFC) syndrome

CFC (cardiofaciocutaneous) syndrome (MIM 115150) has been considered by several authors to be a more severe expression of Noonan syndrome. Affected patients present with congenital heart defects, cutaneous abnormalities, Noonan-like facial features and severe psychomotor developmental delay. We have recently demonstrated that Noonan syndrome can be caused by missense mutations in PTPN11(MIM 176876), a gene that encodes the non-receptor protein tyrosine phosphatase SHP-2. In this report, we have evaluated the possible involvement of mutations in PTPN11 in CFC syndrome. A cohort of 28 CFC subjects rigorously assessed as having CFC based on OMIM diagnostic criteria was examined for mutations in the PTPN11 coding sequence by using DHPLC analysis. The results showed no abnormalities in the coding region of the PTPN11 gene in any CFC patient, nor any evidence of major deletions within the gene suggesting that mutations in other gene(s) are responsible for this syndrome.     

Phenolic compounds and antioxidant properties in the snow alga Chlamydomonas nivalis after exposure to UV light

The snow alga Chlamydomonas nivalis was collected from the Sierra Nevada, California, USA, and examined for its ability to produce phenolic compounds, free proline, and provide antioxidant protection factor in response to UV-A and UV-C light. Exposure of C. nivalis cells to UV-A light (365nm) for 5 days resulted in a 5–12% increase in total phenolics, where as exposure to UV-C light (254 nm) resulted in a 12–24% increase in phenolics after 7 days of exposure. Free proline was not affected by UV-A, but increased markedly after UV-C exposure. A three-fold increase in free proline occurred within two days after exposure to UV-C, but then dropped as cells became bleached. Antioxidant protection factor (PF) increased after treatment of cells with UV-A and remained constant throughout UV-C exposure. Spectral analysis of algal extracts revealed a decrease in absorption in the 215–225 nm region, short-term (2day) stimulation of pigment at 280 nm, and an increase in carotenoids (473 nm), after exposure to UV-A. Snow alga exposed to UV-C light had a different spectrum from that of UV-A exposed cells, i.e. an enhancement of three major peaks at 220, 260, and 280 nm, and loss of absorption in the carotenoid region.We report that UV light exposure, especially in the UV-C range, can stimulate phenolic-antioxidant production in aplanospores of C. nivalis effecting biochemical pathways related to proline metabolism. This revised version was published online in August 2006 with corrections to the Cover Date.     

Simultaneous ethanol production from lactose byKluyveromyces fragilis andZymomonas mobilis

Ethanol production from lactose byKluyveromyces fragilis NRRL 665 in monoculture and coculture with strains ofZymomonas mobilis was studied. One of the strains,Z. mobilis NRRL 1960, when cocultured withK. fragilis, produed 55.2 g/l of ethanol, whereasK. fragilis in monoculture procuded only 36 g/l ethanol from 200 g/l lactose medium. Increased Qp (g ethanol produced/g biomass/h) and Qs (g substrate consumed/g biomass/h) were observed in coculture than in monoculture. However, the residual sugar concentration increased in coculture; this increase might be due to the slow utilization rate of galactose.     

USP14 regulates DNA damage repair by targeting RNF168-dependent ubiquitination

Recent reports have made important revelations, uncovering direct regulation of DNA damage response (DDR)-associated proteins and chromatin ubiquitination (Ubn) by macroautophagy/autophagy. Here, we report a previously unexplored connection between autophagy and DDR, via a deubiquitnase (DUB), USP14. Loss of autophagy in prostate cancer cells led to unrepaired DNA double-strand breaks (DSBs) as indicated by persistent ionizing radiation (IR)-induced foci (IRIF) formation for γH2AFX, and decreased protein levels and IRIF formation for RNF168, an E3-ubiquitin ligase essential for chromatin Ubn and recruitment of critical DDR effector proteins in response to DSBs, including TP53BP1. Consistently, RNF168-associated Ubn signaling and TP53BP1 IRIF formation were reduced in autophagy-deficient cells. An activity assay identified several DUBs, including USP14, which showed higher activity in autophagy-deficient cells. Importantly, inhibiting USP14 could overcome DDR defects in autophagy-deficient cells. USP14 IRIF formation and protein stability were increased in autophagy-deficient cells. Co-immunoprecipitation and colocalization of USP14 with MAP1LC3B and the UBA-domain of SQSTM1 identified USP14 as a substrate of autophagy and SQSTM1. Additionally, USP14 directly interacted with RNF168, which depended on the MIU1 domain of RNF168. These findings identify USP14 as a novel substrate of autophagy and regulation of RNF168-dependent Ubn and TP53BP1 recruitment by USP14 as a critical link between DDR and autophagy. Given the role of Ubn signaling in non-homologous end joining (NHEJ), the major pathway for repair of IR-induced DNA damage, these findings provide unique insights into the link between autophagy, DDR-associated Ubn signaling and NHEJ DNA repair.

Abbreviations: ATG7: autophagy related 7; CQ: chloroquine; DDR: DNA damage response; DUB: deubiquitinase; HR: homologous recombination; IR: ionizing radiation; IRIF: ionizing radiation-induced foci; LAMP2: lysosomal associated membrane protein 2; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MIU1: motif interacting with ubiquitin; NHEJ: non homologous end-joining; PCa: prostate cancer; TP53BP1/53BP1: tumor protein p53 binding protein 1; RNF168: ring finger protein 168; SQSTM1/p62 sequestosome 1; γH2AFX/γH2AX: H2A histone family member X: phosphorylated, UBA: ubiquitin-associated; Ub: ubiquitin; Ubn: ubiquitination; USP14: ubiquitin specific peptidase 14.     

Reduction of hyperhydricity in tissue cultures of oregano (Origanum vulgare) by extracellular polysaccharide isolated from Pseudomonas spp

Hyperhydricity or vitrification is a physiological malformation affecting tissue culture-based propagation of several plant species. A Pseudomonas spp-mediated approach was recently developed to control hyperhydricity in oregano. This bacterium-induced prevention of hyperhydricity helped the establishment of clonal plants in the greenhouse without extensive acclimatization. The prevention of hyperhydricity was specifically linked to mucoid Pseudomonas spp and was characterized by high chlorophyll and reduced water content in oregano shoots. The focus of research reported in this paper was to purify the extracellular mucoid component from Pseudomonas spp and evaluate the effect on hyperhydricity in oregano tissue culture. The extracellular mucoid component was purified by ethanol precipitation. This extracellular mucoid component was confirmed to be a polysaccharide using gas chromatography-mass spectrometry. The effect of purified polysaccharide to prevent or reduce hyperhydricity was tested in oregano clone 0–1. The polysaccharide prevented hyperhydricity in oregano with reduced efficiency compared to bacterial inoculation. This was characterized by higher chlorophyll and reduced water content when compared to uninoculated/untreated oregano shoots. This confirms that the Pseudomonas spp-mediated hyperhydricity reduction in oregano is partially due to its extracellular polysaccharide. This provides a novel approach to develop a media formulation to control hyperhydricity in wide number of plant species where tissue culture is used for clonal propagation.     

Extracellular urease from Arthrobacter creatinolyticus MTCC 5604: scale up,purification and its cytotoxic effect thereof

Molecular Biology Reports - Urease is a potent metalloenzyme with diverse applications. This paper describes the scale up and purification of an extracellular urease from Arthrobacter...     

Microbial degradation of aliphatic and aliphatic-aromatic co-polyesters

Biodegradable plastics (BPs) have attracted much attention since more than a decade because they can easily be degraded by microorganisms in the environment. The development of aliphatic-aromatic co-polyesters has combined excellent mechanical properties with biodegradability and an ideal replacement for the conventional nondegradable thermoplastics. The microorganisms degrading these polyesters are widely distributed in various environments. Although various aliphatic, aromatic, and aliphatic-aromatic co-polyester-degrading microorganisms and their enzymes have been studied and characterized, there are still many groups of microorganisms and enzymes with varying properties awaiting various applications. In this review, we have reported some new microorganisms and their enzymes which could degrade various aliphatic, aromatic, as well as aliphatic-aromatic co-polyesters like poly(butylene succinate) (PBS), poly(butylene succinate)-co-(butylene adipate) (PBSA), poly(ε-caprolactone) (PCL), poly(ethylene succinate) (PES), poly(l-lactic acid) (PLA), poly(3-hydroxybutyrate) and poly(3-hydoxybutyrate-co-3-hydroxyvalterate) (PHB/PHBV), poly(ethylene terephthalate) (PET), poly(butylene terephthalate) (PBT), poly(butylene adipate-co-terephthalate (PBAT), poly(butylene succinate-co-terephthalate) (PBST), and poly(butylene succinate/terephthalate/isophthalate)-co-(lactate) (PBSTIL). The mechanism of degradation of aliphatic as well as aliphatic-aromatic co-polyesters has also been discussed. The degradation ability of microorganisms against various polyesters might be useful for the treatment and recycling of biodegradable wastes or bioremediation of the polyester-contaminated environments.     

Assessment of phenolics-enriched extract and fractions of olive leaves and their antioxidant activities

Recent studies suggest that olive leaf is a significant source of bioactive phenolic compounds comparable to olive oil and fruits. Identifying appropriate extraction methods is thus an important step to increase the yield of such bioactive components from olive leaf, which is otherwise agricultural waste. The present study evaluates phenolic contents and compositions of olive leaf extracted by several solvent methods and to further establish their antioxidant activities using various radical scavenging systems. Total flavonoid and phenolic contents were significantly higher in the 80% ethanol extract, butanol, and ethylacetate fractions than hexane, chloroform and water fractions (p < 0.05). Oleuropein was identified as a major phenolic compound with considerable contents in these major three fractions and the extract that correlated with their higher antioxidant and radical scavenging. These results indicate that olive leaf contains significant amounts of oleuropein and phenolics, important factors for antioxidant capacity, which can be substantially modified by different extraction methods.     

Deterioration of semen quality and sperm-DNA integrity as influenced by cigarette smoking in fertile and infertile human male smokers—A prospective study

In modernized lifestyle smoking is one of the trendy, psychological, and socioeconomic scenarios of young adolescents mainly in the age of the reproductive stage. Based on a number of cigarettes smoked, age, and duration of the smoke, the study aims to search for the profound effects of smoking and its impact on semen parameters, sperm-DNA integrity, and fragmentation of sperm DNA with cotinine and apoptotic caspase-3 marker in the seminal plasma of fertile and infertile smokers. To determine oxidative damage by 8-hydroxy deoxyguanosine (8-OHdG) from isolated sperm DNA (steps: reactive oxygen species washing by nitro blue tetrazolium (NBT), sperm lysis, salt digestion, ethanol washing, and finally with high-performance liquid chromatography analysis). Level of DNA fragmentation (percentage) in native and intact DNA, the activity of caspase-3 in infertile smokers will be compared with the control group of nonsmokers. Also, the sperm viability was visualized by eosin-nigrosin and aniline blue staining. Cotinine is one of the best markers of smoking. The cotinine level (2224.24 ± 1.19 *** ng/mL), when abundant it negative correlates with morphology and rapid motility in infertile smokers than nonsmokers. Gel preprogram measured the sperm integrity and was found to be less in smokers than nonsmokers. The spermatic oxidative marker 8-OHdG was high and gave an R ² value of 0.9104 with morphology and 0.9007 for rapid motility of infertile sperm, respectively. Infertile smoking subjects (<10 cigarettes/day) had significant changes increase in sperm fragmentation, caspase-3, and cotinine while negative impact with motility, morphology, and pH of semen compared with fertile, infertile nonsmoking subjects.