Anatomical and physiological evidence for polarisation vision in the nocturnal bee <Emphasis Type="Italic">Megalopta genalis</Emphasis>

The presence of a specialised dorsal rim area with an ability to detect the e-vector orientation of polarised light is shown for the first time in a nocturnal hymenopteran. The dorsal rim area of the halictid bee Megalopta genalis features a number of characteristic anatomical specialisations including an increased rhabdom diameter and a lack of primary screening pigments. Optically, these specialisations result in wide spatial receptive fields (Δρ = 14°), a common adaptation found in the dorsal rim areas of insects used to filter out interfering effects (i.e. clouds) from the sky. In this specialised eye region all nine photoreceptors contribute their microvilli to the entire length of the ommatidia. These orthogonally directed microvilli are anatomically arranged in an almost linear, anterior–posterior orientation. Intracellular recordings within the dorsal rim area show very high polarisation sensitivity and a sensitivity peak within the ultraviolet part of the spectrum.     

Conversion of Unprotected Amino-Link Oligonucleotides into Their Tetramethylguanidinium Derivatives

Abstract

The preparation of tetramethylguanidinium oligodeoxynucleotide (ODN) derivatives by reaction of the corresponding aminoalkyl-ODN with the uronium salts HBTU, TBTU or HATU, respectively, is described. The binding affinity of the new tetramethylguanidinium ODN derivatives was determined.     

Nasitrema sp.-associated encephalitis in a striped dolphin (Stenella coeruleoalba) stranded in the Gulf of Mexico.

An immature female striped dolphin (Stenella coeruleoalba) found dead on a northwestern Florida beach in 1988 exhibited severe inflammation bilaterally in the dorsal and mid-thalamus in association with adult trematodes (Nasitrema sp.) and trematode eggs. Numerous specimens of Nasitrema sp. also were present in the pterygoid sinuses. Pneumonia in association with a heavy growth of Vibrio damsela was observed also. This report confirms the occurrence of Nasitrema sp.-associated encephalitis in striped dolphins and in small cetaceans from the Gulf of Mexico.     

An efficient method for the extraction of DNA from formalin-fixed, paraffin-embedded tissue by sonication

A method was developed for fast and efficient isolation of DNA from formalin-fixed, paraffin-embedded tissue sections for subsequent use in PCRs and DNA hybridization assays. The method relies on the use of a sonicating water bath to disrupt tissue samples to which a small amount of micro-sized glass beads have been added. The sonicating glass beads provide fast and efficient physical shearing of fixed tissue sections, allowing for quick release and solubilization of the DNA. The extraction process from paraffin section to amplifiable target DNA takes 30 minutes. The method eliminates the need for repetitive solvent extractions and exhaustive proteinase K digestion. PCR amplification of human genomic and viral target sequences was successfully carried out on DNA isolated from a number of different types of normal and infected tissues.     

Mechanism of metolachlor action due to alterations in cell cycle progression

Metolachlor, a commonly used herbicide in the Midwestern USA, functions by inhibiting chlorophyll and protein synthesis in target plants. Herbicide exposure has led to detrimental effects in several organisms, affecting their growth and behavior; however, its mechanism of action in nontarget organisms is not yet clear. The EPA does not currently have enforceable regulations for maximal limits allowed in drinking water. Previous growth studies from our lab have demonstrated that increasing metolachlor concentrations and increasing time of exposure results in decreased growth of liver cells. The objective of this study was to elucidate a mechanism for this decrease of HepG2 cell growth after herbicide exposure. Results show that metolachlor at environmentally relevant levels (50–100 ppb) that previously led to decreased cell number does not lead to cell death by either necrosis or apoptosis. However, it was demonstrated that the levels of the retinoblastoma protein including two of its hyperphosphorylated forms are decreased in metolachlor exposed cells possibly leading to cell cycle arrest. The levels of another protein involved in cell cycle progression, p53, a mediator in the DNA damage response of cells, was not significantly altered except at the highest level of metolachlor (1,000 ppb) and after a 72-h exposure. These results suggest that the decrease in cell number after low-level metolachlor exposure is most likely due to an alteration in the cell cycle and not due to cell death in human liver cells.     

Tissue distribution of a plasmid DNA encoding Hsp65 gene is dependent on the dose administered through intramuscular delivery

In order to assess a new strategy of DNA vaccine for a more complete understanding of its action in immune response, it is important to determine the in vivo biodistribution fate and antigen expression. In previous studies, our group focused on the prophylactic and therapeutic use of a plasmid DNA encoding the Mycobacterium leprae 65-kDa heat shock protein (Hsp65) and achieved an efficient immune response induction as well as protection against virulent M. tuberculosis challenge. In the present study, we examined in vivo tissue distribution of naked DNA-Hsp65 vaccine, the Hsp65 message, genome integration and methylation status of plasmid DNA. The DNA-Hsp65 was detectable in several tissue types, indicating that DNA-Hsp65 disseminates widely throughout the body. The biodistribution was dose-dependent. In contrast, RT-PCR detected the Hsp65 message for at least 15 days in muscle or liver tissue from immunized mice. We also analyzed the methylation status and integration of the injected plasmid DNA into the host cellular genome. The bacterial methylation pattern persisted for at least 6 months, indicating that the plasmid DNA-Hsp65 does not replicate in mammalian tissue, and Southern blot analysis showed that plasmid DNA was not integrated. These results have important implications for the use of DNA-Hsp65 vaccine in a clinical setting and open new perspectives for DNA vaccines and new considerations about the inoculation site and delivery system.     

Kinetics, substrate specificity, and stereospecificity of two new protein tyrosine phosphatase-like inositol polyphosphatases from Selenomonas lacticifex

Inositol polyphosphatases (IPPases) play an important role in the metabolism of inositol polyphosphates, a class of molecules involved in signal transduction. Here we characterize 2 new protein tyrosine phosphatase-like IPPases (PhyAsl and PhyBsl) cloned from Selenomonas lacticifex that can hydrolyze myo-inositol hexakisphosphate (InsP6) in vitro. To determine their preferred substrates and stereospecificity of InsP6 dephosphorylation, a combination of kinetic and high-performance ion pair chromatography studies were conducted. Despite only 33% amino acid sequence identity between them, both enzymes display strict specificity for IPP substrates and cleave InsP6 primarily at the d-3-phosphate position (>90%). Furthermore, both enzymes predominantly degrade InsP6 to Ins(2)P via identical and very specific routes of dephosphorylation (3,4,5,6,1). Despite these similarities, PhylAsl is shown to have a slight kinetic preference for the major inositol pentakisphosphate intermediate in its InsP6 hydrolysis pathway, whereas PhyBsl displays a unique and substantial preference for an inositol tetrakisphosphate intermediate.