Expression of two methionine-rich storage protein genes of Plutella xylostella (L.) in response to development, juvenile hormone-analog and pyrethroid

We cloned and characterized cDNA of two storage protein (SP) genes, PxSP1 and PxSP2, from the diamondback moth, Plutella xylostella (Lepidoptera: Yponomeutidae) and investigated their expression. PxSP1 and PxSP2 each encoded a putative protein of 91 kDa. Nucleotide and deduced amino acid identities between the two genes were 79% and 82%, respectively. Amino acid composition (methionine>4%), sequence homology with other insect storage proteins and the phylogenetic analysis suggested that the genes belong to the subfamily of moderately methionine-rich SP genes. The genes were predominantly expressed in the last instar female larvae and the mRNA levels were suppressed by treatment with a juvenile hormone-analog. Treatment of female larvae with sublethal dose of a pyrethroid caused a significant increase in mRNA levels of both genes. Induction of PxSP1 and PxSP2 genes as a result of pyrethroid application may have implications with respect to reproduction as methionine-rich proteins are known as a key element for egg production.     

Tunable Salisbury Screen Absorber Using Square Lattice of Plasmonic Nanodisk

We propose a novel polarization independent Salisbury screen absorber to provide tunable resonant absorption at terahertz (THz) frequencies. The Salisbury screen absorber is designed by using a planar array of thin gold nanodisks arranged in a square lattice. Certain configurations of Salisbury screen have multiple distinctive absorption bands that support near-unity/FWHM absorption bandwidth reaching 36 THz/169 THz, respectively. Moreover, the absorption bandwidth depends upon the optical thickness of the dielectric spacer between the metasurface and the metallic ground plane. The proposed tunable Salisbury screen absorber can find practical applications in photonic detection, imaging, sensing, and solar cells at optical frequencies.     

Neuronal Calcium Sensor-1 Binds the D2 Dopamine Receptor and G-protein-coupled Receptor Kinase 1 (GRK1) Peptides Using Different Modes of Interactions

Neuronal calcium sensor-1 (NCS-1) is the primordial member of the neuronal calcium sensor family of EF-hand Ca²⁺-binding proteins. It interacts with both the G-protein-coupled receptor (GPCR) dopamine D2 receptor (D2R), regulating its internalization and surface expression, and the cognate kinases GRK1 and GRK2. Determination of the crystal structures of Ca²⁺/NCS-1 alone and in complex with peptides derived from D2R and GRK1 reveals that the differential recognition is facilitated by the conformational flexibility of the C-lobe-binding site. We find that two copies of the D2R peptide bind within the hydrophobic crevice on Ca²⁺/NCS-1, but only one copy of the GRK1 peptide binds. The different binding modes are made possible by the C-lobe-binding site of NCS-1, which adopts alternative conformations in each complex. C-terminal residues Ser-178–Val-190 act in concert with the flexible EF3/EF4 loop region to effectively form different peptide-binding sites. In the Ca²⁺/NCS-1·D2R peptide complex, the C-terminal region adopts a 3₁₀ helix-turn-3₁₀ helix, whereas in the GRK1 peptide complex it forms an α-helix. Removal of Ser-178–Val-190 generated a C-terminal truncation mutant that formed a dimer, indicating that the NCS-1 C-terminal region prevents NCS-1 oligomerization. We propose that the flexible nature of the C-terminal region is essential to allow it to modulate its protein-binding sites and adapt its conformation to accommodate both ligands. This appears to be driven by the variability of the conformation of the C-lobe-binding site, which has ramifications for the target specificity and diversity of NCS-1.     

Identification of Recombination in the Envelope Gene of Simian Foamy Virus Serotype 2 Isolated from Macaca cyclopis

The full-length sequence of simian foamy virus serotype 2 (SFVmcy-2), isolated from a Taiwanese macaque, was determined. SFVmcy-2 was highly related to SFV serotype 1 (SFVmcy-1), an isolate from the same species, except in the putative receptor binding domain (RBD) in env, which contained novel sequences related to SFV serotype 3 (SFVagm-3), isolated from an African green monkey. The results identify a potential region of neutralization in SFVs and demonstrate recombination between genetically divergent foamy viruses.     

Oxazolones: new tyrosinase inhibitors; synthesis and their structure-activity relationships

The tyrosinase inhibitory potential of seventeen synthesized oxazolone derivatives has been evaluated and their structure-activity relationships developed in the present work. All the synthesized derivatives, 3-19, demonstrated excellent in vitro tyrosinase inhibitory properties having IC50 values in the range of 1.23+/-0.37-17.73+/-2.69 microM, whereas standard inhibitors l-mimosine and kojic acid have IC50 values 3.68+/-0.02 and 16.67+/-0.52 microM,, respectively. Compounds 4-8 having IC50 values 3.11+/-0.95, 3.51+/-0.25, 3.23+/-0.66, 1.23 +/- 0.37, and 2.15+/-0.75, respectively, were found to be very active members of the series, even better than both the standard inhibitors. However, compounds 3, 9-11, 13, 14, 16, 17, and 19 were found to be better than kojic acid but not l-mimosine. (2-Methyl-4-[E,2Z)-3-phenyl-2-propenyliden]-1,3-oxazol-5(4H)-one (7) bearing a cinnamyol residue at C-4 of oxazolone moiety and an IC50 = 1.23+/-0.37 microM was found to be the most active one among all tested compounds. These studies reveal that the substitution of functional group (s) at C-4 and C-2 positions plays a vital role in the activity of this series of compounds. It is concluded that compound 7 may act as a potential lead molecule to develop new drugs for the treatment of tyrosinase based disorders.     

Evidence for regulated interleukin-4 expression in chondrocyte-scaffolds under in vitro inflammatory conditions

Objective

To elucidate the anti-inflammatory and anabolic effects of regulated expression of IL-4 in chondrocyte-scaffolds under in vitro inflammatory conditions.

Methods

Mature articular chondrocytes from dogs (n = 3) were conditioned through transient transfection using pcDNA3.1.cIL-4 (constitutive) or pCOX-2.cIL-4 (cytokine-responsive) plasmids. Conditioned cells were seeded in alginate microspheres and rat-tail collagen type I matrix (CaReS®) to generate two types of tissue-engineered 3-dimensional scaffolds. Inflammatory arthritis was simulated in the packed chondrocytes through exogenous addition of recombinant canine (rc) IL-1β (100 ng/ml) plus rcTNFα (50 ng/ml) in culture media for 96 hours. Harvested cells and culture media were analyzed by various assays to monitor the anti-inflammatory and regenerative (anabolic) properties of cIL-4.

Results

cIL-4 was expressed from COX-2 promoter exclusively on the addition of rcIL-1β and rcTNFα while its expression from CMV promoter was constitutive. The expressed cIL-4 downregulated the mRNA expression of IL-1β, TNFα, IL-6, iNOS and COX-2 in the cells and inhibited the production of NO and PGE₂ in culture media. At the same time, it up-regulated the expression of IGF-1, IL-1ra, COL2a1 and aggrecan in conditioned chondrocytes in both scaffolds along with a diminished release of total collagen and sGAG into the culture media. An increased amount of cIL-4 protein was detected both in chondrocyte cell lysate and in concentrated culture media. Neutralizing anti-cIL-4 antibody assay confirmed that the anti-inflammatory and regenerative effects seen are exclusively driven by cIL-4. There was a restricted expression of IL-4 under COX-2 promoter possibly due to negative feedback loop while it was over-expressed under CMV promoter (undesirable). Furthermore, the anti-inflammatory /anabolic outcomes from both scaffolds were reproducible and the therapeutic effects of cIL-4 were both scaffold- and promoter-independent.

Conclusions

Regulated expression of therapeutic candidate gene(s) coupled with suitable scaffold(s) could potentially serve as a useful tissue-engineering tool to devise future treatment strategies for osteoarthritis.     

Exogenously applied glycinebetaine enhances seed and seed oil quality of maize (Zea mays L.) under water deficit conditions

A field trial was carried out to appraise up to what extent exogenous application of a potential osmoprotectant, glycinebetaine (GB), could ameliorate the inhibitory effects of shortage of water on maize seed and seed oil composition and oil antioxidant potential. Two maize cultivars, Agaiti-2002 (drought tolerant) and EV-1098 (drought sensitive), were exposed to drought treatments at the vegetative growth stage. Both the maize cultivars used in the present study are being widely cultivated in Pakistan and have been an important source of developing different maize hybrids. Two levels of glycinebetaine (0 or 30 mM) were foliar-applied at the vegetative stage. Water stress reduced the kernel sugar, oil, protein, moisture contents and most of the seed micro- and macro-nutrients analyzed of both maize cultivars, but it increased the contents of seed fiber and ash contents. Among different seed oil un-saturated fatty acids, water stress increased the oil oleic acid contents with a decrease in linoleic acid contents, which resulted in increased oil oleic/linoleic ratio of both maize cultivars. However, no variation was observed in oil stearic and palmitic acid contents due to water stress. A considerable increase in seed oil α-, γ-, δ- and total tocopherols and flavonoids was observed in both maize cultivars. However, oil phenolic content and 1,1′-diphenyl-2-picryl-hydrazyl (DPPH) free radical scavenging activity decreased. Foliar-applied GB significantly increased the contents of seed sugar, oil, protein, moisture, fiber, ash, GB contents and micro- and macro-nutrients of both maize cultivars under well irrigated and water deficit conditions. Furthermore, exogenous application of GB increased the oil oleic and linoleinic acid contents. All different lipophilic compounds estimated in the seed oil increased due to foliar applied GB. Furthermore, GB also increased seed oil antioxidant activity appraised in terms of oil DPPH free radical scavenging activity. By summarizing the results, it seemed that exogenously applied GB remained in intact form until later stages of growth and counteracted the inhibitory effects of water deficit on seed and seed oil composition similarly of both maize cultivars.     

Effects of Plant Growth Regulators on Seed Filling,Endogenous Hormone Contents and Maize Production in Semiarid Regions

In this study, we determined whether the application of uniconazole alone or combined with ethephon could enhance the seed-filling rates in maize by regulating the endogenous hormone contents. Uniconazole was applied to the foliage at concentrations of 0 (CK), 25 (U₂₅), 50 (U₅₀) and 75 (U₇₅) mg L⁻¹ at the 12-leaf stage. In addition, uniconazole was applied to the foliage at the 12-leaf stage and ethephon at 10 days after silking stage at concentrations of 0 (CK), 25 mg L⁻¹ uniconazole + 100 mg L⁻¹ ethephon (U₂₅ + E₁₀₀), 50 mg L⁻¹ uniconazole + 200 mg L⁻¹ ethephon (U₅₀ + E₂₀₀) and 75 mg L⁻¹ uniconazole + 300 mg L⁻¹ ethephon (U₇₅ + E₃₀₀). Uniconazole applied alone or in combination with ethephon significantly improved ear characters and grain yield. Uniconazole applied alone or combination with ethephon significantly improved the dry matter accumulation in seeds and seed-filling rates. Uniconazole significantly increased the abscisic acid (ABA) and zeatin (Z) + zeatin riboside (ZR) contents of seeds, but reduced the gibberellic acid (GA) contents. The application of uniconazole combined with ethephon decreased the ABA, Z + ZR and GA contents in seeds. The ABA and Z + ZR contents were significantly positively correlated, whereas the GA content was negatively correlated with the maximum seed weight, maximum seed-filling rate and mean seed-filling rate. The application of uniconazole alone significantly improved the seed-filling rates in maize by regulating the endogenous hormone contents. Thus, we conclude that uniconazole application at 50 mg L⁻¹ in the 12-leaf stage can enhance the maize production.

     

Role of ACE and PAI-1 Polymorphisms in the Development and Progression of Diabetic Retinopathy

In the present study we determined the association of angiotensin converting enzyme (ACE) and plasminogen activator inhibitor-1 (PAI-1) gene polymorphisms with diabetic retinopathy (DR) and its sub-clinical classes in Pakistani type 2 diabetic patients. A total of 353 diabetic subjects including 160 DR and 193 diabetic non retinopathy (DNR) as well as 198 healthy controls were genotyped by allele specific polymerase chain reaction (PCR) for ACE Insertion/Deletion (ID) polymorphism, rs4646994 in intron 16 and PAI-1 4G/5G (deletion/insertion) polymorphism, rs1799768 in promoter region of the gene. To statistically assess the genotype-phenotype association, multivariate logistic regression analysis was applied to the genotype data of DR, DNR and control individuals as well as the subtypes of DR. The ACE genotype ID was found to be significantly associated with DR (p = 0.009, odds ratio (OR) 1.870 [95% confidence interval (CI) = 1.04–3.36]) and its sub-clinical class non-proliferative DR (NPDR) (p = 0.006, OR 2.250 [95% CI = 1.098–4.620]), while PAI polymorphism did not show any association with DR in the current cohort. In conclusion in Pakistani population the ACE ID polymorphism was observed to be significantly associated with DR and NPDR, but not with the severe form of the disease i.e. proliferative DR (PDR).