The chemical compound ‘Heatin’ stimulates hypocotyl elongation and interferes with the Arabidopsis NIT1-subfamily of nitrilases

Temperature passively affects biological processes involved in plant growth. Therefore, it is challenging to study the dedicated temperature signalling pathways that orchestrate thermomorphogenesis, a suite of elongation growth-based adaptations that enhance leaf-cooling capacity. We screened a chemical library for compounds that restored hypocotyl elongation in the pif4-2–deficient mutant background at warm temperature conditions in Arabidopsis thaliana to identify modulators of thermomorphogenesis. The small aromatic compound ‘Heatin’, containing 1-iminomethyl-2-naphthol as a pharmacophore, was selected as an enhancer of elongation growth. We show that ARABIDOPSIS ALDEHYDE OXIDASES redundantly contribute to Heatin-mediated hypocotyl elongation. Following a chemical proteomics approach, the members of the NITRILASE1-subfamily of auxin biosynthesis enzymes were identified among the molecular targets of Heatin. Our data reveal that nitrilases are involved in promotion of hypocotyl elongation in response to high temperature and Heatin-mediated hypocotyl elongation requires the NITRILASE1-subfamily members, NIT1 and NIT2. Heatin inhibits NIT1-subfamily enzymatic activity in vitro and the application of Heatin accordingly results in the accumulation of NIT1-subfamily substrate indole-3-acetonitrile in vivo. However, levels of the NIT1-subfamily product, bioactive auxin (indole-3-acetic acid), were also significantly increased. It is likely that the stimulation of hypocotyl elongation by Heatin might be independent of its observed interaction with NITRILASE1-subfamily members. However, nitrilases may contribute to the Heatin response by stimulating indole-3-acetic acid biosynthesis in an indirect way. Heatin and its functional analogues present novel chemical entities for studying auxin biology.     

Genetic and molecular analysis of Arabidopsis thaliana (ecotype ‘Estland’) transformed with Agrobacterium

Summary Agrobacterium-mediated transformation of Arabidopsis, ecotype ‘Estland’, was established from root explants using kanamycin selection. Continuous light during callus and shoot induction phases was promotive for shoot regeneration, as compared to light/dark cycles. Use of optimized conditions for transformation led to the formation of kanamycin-resistant calluses (up to 77%) and transformed plantlets at a frequency of up to 45%. Southern analysis showed the presence of 1.2. or more T-DNA inserts in 33%, 50%, and 17% of the primary transformants, respectively. Mendelian, as well as non-Mendelian, inheritance patterns were obtained upon screening the progeny (T1) of various transformants for the expression of gus and nptII genes; the analysis of some of these transformants at the molecular level also corroborated the Mendelian inheritance pattern. Moreover, genotypes of the T1 progeny could be predicted on the basis of T2 progeny analysis.     

A New Enzyme Immunoassay for the Quantitative Determination of Classical Autotaxins (ATXα, ATXβ, and ATXγ) and Novel Autotaxins (ATXδ and ATXε)

Background

Autotaxin (ATX) is a secreted enzyme that converts lysophosphatidylcholine to lysophosphatidic acid, a potent bioactive lipid mediator, through its lysophospholipase D activity. Although five alternative splicing isoforms of ATX have been identified as ATXα, ATXβ, ATXγ, ATXδ, and ATXε and the expression patterns of each isoform differ among several tissues, the clinical significance of each isoform remains to be elucidated.

Methods

Anti-ATXβ and anti-ATXδ monoclonal antibodies were produced by immunization with recombinant human ATXβ and ATXδ expressed using a baculovirus system, respectively. We then developed enzyme immunoassays to measure the serum concentrations of “classical ATX” (ATXα, ATXβ, and ATXγ) and “novel ATX” (ATXδ and ATXε) antigens and evaluated the usefulness of these assays using human serum samples.

Results

The with-run and between-run precision, interference, detection limit, and linearity studies for the present assay were well validated. In healthy subjects, the serum concentrations of classical ATX and novel ATX were significantly (P < 0.01) higher in women than in men, while the ratios of classical ATX or novel ATX to total ATX were not different between women and men. The concentrations of both classical ATX and novel ATX in normal pregnant subjects and patients with chronic liver diseases or follicular lymphoma were significantly higher than those in healthy subjects, while the ratio of both ATX isoforms to total ATX did not vary among these groups.

Conclusions

We have developed a new enzyme immunoassay to determine the concentrations of classical ATX and novel ATX in human serum. These assays may be helpful for elucidating the distinct functional roles of each ATX isoform, which are largely unknown at present.     

Arabidopsis thaliana ‘extra-large GTP-binding protein’ (AtXLG1): a new class of G-protein

Heterotrimeric GTP-binding proteins, composed of , , and subunits, are involved in signal transduction pathways in animal and plant systems. In plants, physiological analyses implicate heterotrimeric G-proteins in ion channel regulation, light signaling, and hormone and pathogen responses. However, only one class of plant G genes has been identified to date. We have cloned a novel gene, Arabidopsis thaliana extra-large GTP-binding protein (AtXLG1). AtXLG1 appears to be a member of a small gene family and is transcribed in all tissues assayed: roots, leaves, stems, flowers, and fruits. The conceptually translated protein from AtXLG1 is 99 kDa, twice as large as typical G proteins. The carboxy-terminal half of the AtXLG1 protein has significant homology to animal and plant G proteins. This region includes a GTP-binding domain, a predicted helical domain, and an aspartate/glutamate-rich loop, which are characteristics of G's. Despite the absence of some of the amino acids implicated in GTP binding and hydrolysis by crystallographic and mutational analyses of mammalian G's, recombinant AtXLGl binds GTP with specificity. The amino-terminal region of AtXLGl contains domains homologous to the bacterial TonB-box, which is involved in energy transduction between the inner and outer bacterial membranes, and to zinc-finger proteins. Given the unique structure of AtXLG1, it will be of interest to uncover its physiological functions.     

Don't Blame the ‘Bio’ — Blame the ‘Ethics’: Varieties of (bio) ethics and the challenge of pluralism

We tend to think that the difficulties in bioethics spring from the novel and alarming issues that arise due to discoveries in the new biosciences and biotechnologies. But many of the crucial difficulties in bioethics arise from the assumption we make about ethics. This paper offers a brief overview of bioethics, and relates ethical 'principlism' to 'ethical fundamentalism.' It then reviews some alternative approaches that have emerged during the second phase of bioethics and argues for a neo-Aristotelian approach. Misconceptions about ethical principles and ethical reasoning not only distort our views of the business of bioethics, but they also prevent us from facing up to the formidable problems posed by ethical pluralism in so-called liberal societies.     

‘Scooty Girls’: Mobility and Intimacy at the Margins of Kathmandu

ABSTRACT

The recent explosion in the number of scooters on the road in the Kathmandu Valley signifies a transformation of women's mobility. Scooters allow women to move from the back of a motorbike to the driver's seat. In a context in which most women were discouraged from leaving the house without a reason, the ability to drive oneself rather than call upon a male escort has pushed the gendered boundaries of acceptable behaviour for young women outwards, metaphorically and literally. This paper analyses young, unmarried women's newfound mobility and their capacity to drive to peripheral, ‘out-of-the-way’ places that offer respite from the bustle and pollution of the city, scenic views of the valley below, and something rarely achieved in the past – privacy. Scooters offer a way out of the policed realm of a young woman's home and neighbourhood, leading to exploration and a new space for the creation of intimacy.     

Restriction fragment analysis of ‘null’ forms at the Gli-1 loci of bread and durum wheats

Summary Wheat accessions lacking some of the - and -gliadin components encoded by the Gli-1 loci on the short arm of chromosome 1D in bread wheat and chromosome 1A in durum wheat were studied by two-dimensional polyacrylamide gel electrophoresis and restriction fragment analysis. Digested genomic DNAs of normal and null forms were probed with a cDNA clone related to -/-gliadins and with a genomic clone encoding an LMW subunit of glutenin. The hybridisation patterns with the -/-gliadin probe were similar to those of cvs Chinese Spring and Langdon used as standards for bread and durum wheats, respectively, but several restriction fragments located on the 1D chromosome of bread wheat and the 1A chromosome of durum wheat were absent in the null forms. In addition, specific LMW glutenin fragments encoded by the same chromosomes were also absent in the null forms, suggesting that simultaneous deletions of blocks of genes for both -/-gliadins and LMW glutenins had occurred. Comparisons of the protein and RFLP patterns enabled some proteins to be mapped to specific restriction fragments.     

‘… The End of the Beginning’: Cdk1 Thresholds and Exit from Mitosis

Exit from mitosis requires the proteolytic degradation of mitotic cyclins, which is instigated by the APC/C ubiquitin ligase. The coincidence of mitotic cyclin B1 degradation with the onset of anaphase intuitively suggested a requirement of cyclin degradation for sister chromatid separation. While this hypothesis has originally been refuted, evidence that cyclin B1 degradation is required for anaphase during meiosis has been obtained, while its requirement for anaphase during mitosis is still more controversial. By studying human cells engineered to express non-degradable cyclin B1, we have recently shown that stable cyclin B1 affects progression through mitosis at various steps in a dose-dependent manner. These experiments suggest that controlled exit from mitosis might involve CDK activity thresholds for important late mitotic events, such as the onset of anaphase, formation of the spindle midzone, the onset of cytokinesis, cellular abscission and chromosome decondensation.     

Variant detection at the δ opioid receptor (OPRD1) locus and population genetics of a novel variant affecting protein sequence

The three opioid receptor genes, and in particular the mu and delta loci (OPRM1 and OPRD1, respectively), are compelling candidates to influence risk for substance dependence. Previous study of a variant at the OPRD1 locus, T921C, has shown association with opioid dependence. This variant does not alter protein sequence, and could not be directly responsible for a physiologic effect. We sequenced the OPRD1 coding region in six individuals with differing T921C alleles, to identify new common variants more likely to explain the association with phenotype. We identified one novel variant in exon 1, 80T-->G, which predicts a change in amino acid sequence from phenylalanine (80T) to cysteine (80G) (F27C). We present here basic population genetics of this variant, and population genetic data for the T921C variant. We found significant differences in allele frequency between populations, and a maximum frequency of the 80G allele of 9%, in each of two European populations. This variant could contribute to the previously reported association results.     

The age of immigration of the vertebrate faunas found at Gargano (Apulia,Italy) and Scontrone (l’Aquila,Italy)

Recently a discussion is taking place about the Scontrone (l’Aquila) and Gargano (Apulia, Italy) mammal faunas and the age of their immigration. Mazza and Rustioni (2008) dated the Scontrone mammal fossils as Tortonian on the basis of their position in the Lithothamnium Limestone and came to the conclusion that some elements of the Scontrone and Gargano faunas must have colonised the area in Oligocene or Early Miocene times. Van den Hoek Ostende et al. (2009) disagreed with this interpretation and suggested a Late Miocene (10 Ma) age for the time of immigration. We think the arguments to place Scontrone in the Tortonian are not convincing. An analysis of the potential ancestors of each of the Gargano faunal components shows that a Messinian age for the immigration is fully compatible with the distribution of these ancestors in the European Miocene.     

The fertility of women Poland in the period of trasformation of the political and economics system (the 80’s and 90’s)

Wielkopolska (western Poland) is a region, with good reason, called the main food base of Poland. All indices of agricultural production have always been there (since 19th century) much higher than elsewhere in the country (Ratajczak 1977/1978, Buszko 1984, Gorzelak 1989, Czyzewski et al. 1992; table 1). Such a situation had a beneficial effect on the amount of the income generated by a family of Wielkopolska and, thus, decided on its better financial status. Additionally, rural inhabitants of this region have always been characterised by a more advantageous educational structure in comparison with people of other Polish villages. Generally, it is easy to detect the relationship between the life standard, the level of education and family planning. The aim of this paper is to assess fertility of women from villages of Wielkopolska in the period of transformation of the political and economic system in Poland.     

The ‘Frank J. Myers Rotifera Collection’ at the Academy of Natural Sciences of Philadelphia

The Academy of Natural Sciences of Philadelphia (ANSP) houses the world’s most comprehensive collection of Rotifera on microscope slides. A particular strength of this major reference collection and type repository lies in the excellent preparation of specimens in life-like extended state and its balanced coverage of the phylum. The collection is almost worldwide in scope, and currently comprises 774 valid taxa, equalling a taxon coverage of 88.2% (families), 75.4% (genera), and 39.1% (species) compared to taxon numbers currently known to occur worldwide. A searchable database plus illustrated catalog has now been published on CD-ROM and on the Academy’s website. The poor representation of Rotifera in natural history collections and their generally poor curation and conservation is commented upon.     

The mineral composition of the tests of ‘testate amoebae’ (Amoebozoa,Arcellinida): The relative importance of grain availability and grain selection

Nine peatlands were selected according to their various geological setting in the eastern part of France. The diversity of mineral particles, including xenosomes (agglutinated particles) and idiosomes (secreted particles), were analysed, together with associated morphological characteristics, for 7 species of ‘testate amoeba’ (order Arcellinida). The combined use of an Environmental Scanning Electron Microscope equipped with an EDS device and microprobe analyses is suitable for conducting an elemental analysis and subsequently enabled the determination of 24 different minerals. Such mineral diversity has never been recorded before. We conclude that the testate amoebae select grains according to their size from those available within their immediate environment. Availability in turn reflects the geological surroundings and the stability of the different kinds of grains, while their size seems to be a function of the distance from the source.     

Coverage and Quality of Antenatal Care Provided at Primary Health Care Facilities in the ‘Punjab’ Province of ‘Pakistan’

Background

Antenatal care is a very important component of maternal health services. It provides the opportunity to learn about risks associated with pregnancy and guides to plan the place of deliveries thereby preventing maternal and infant morbidity and mortality. In ‘Pakistan’ antenatal services to rural population are being provided through a network of primary health care facilities designated as ''Basic Health Units and Rural Health Centers. Pakistan is a developing country, consisting of four provinces and federally administered areas. Each province is administratively subdivided in to ‘Divisions’ and ‘Districts’. By population ‘Punjab’ is the largest province of Pakistan having 36 districts. This study was conducted to assess the coverage and quality antenatal care in the primary health care facilities in ‘Punjab’ province of ‘Pakistan’.

Methods

Quantitative and Qualitative methods were used to collect data. Using multistage sampling technique nine out of thirty six districts were selected and 19 primary health care facilities of public sector (seventeen Basic Health Units and two Rural Health Centers were randomly selected from each district. Focus group discussions and in-depth interviews were conducted with clients, providers and health managers.

Results

The overall enrollment for antenatal checkup was 55.9% and drop out was 32.9% in subsequent visits. The quality of services regarding assessment, treatment and counseling was extremely poor. The reasons for low coverage and quality were the distant location of facilities, deficiency of facility resources, indifferent attitude and non availability of the staff. Moreover, lack of client awareness about importance of antenatal care and self empowerment for decision making to seek care were also responsible for low coverage.

Conclusion

The coverage and quality of the antenatal care services in ‘Punjab’ are extremely compromised. Only half of the expected pregnancies are enrolled and out of those 1/3 drop out in follow-up visits.     

The roles of the Chantransia phase of Lemanea (Lemaneaceae,Batrachospermales, Rhodophyta) and of the ‘Mushroom’ phase of Himanthalia (Himanthaliaceae,Fucales, Phaeophyta)

Summary

The life cycle of the Batrachospermales (freshwater florideophyte red such as Batrachospermum, Lemanea) is a shortened variant of the ‘normal’ marine florideophyte life cycle. The perennial Chantransia diploid phase is microscopic and encrusting. Each winter it produces one or more semi-erect haploid gametophytes by vegetative meiosis. Gamete production and fertilization is followed by production of diploid carposporophytes; these produce diploid carpospores which disperse, and regenerate the Chantransia phase. The question of the extent to which the Chantransia phase contributes resources to the gametophyte was approached by physiological-hydrodynamic modelling. These computations suggest that the photosynthetic rates in situ on an area basis are 20 times greater for the gametophyte than the Chantransia phase; this agrees with the observed ratios of peak biomass. The conclusion is that the Chantransia phase has a negligible role in provisioning the growing gametophyte, and that the role of the Chantransia phase is to occupy space with living biomass throughout the year, including exposure at summer drawdown, and (perhaps) by dispersal via production of monospores. A similar conclusion is arrived at on the basis of biomass data for the role of the perennial ‘mushroom’ phase of the semelparous marine Fucalean brown alga Himanthalia elongata in relation to the short-lived but much larger reproductive receptacles.