DNA hypermethylation in Drosophila melanogaster causes irregular chromosome condensation and dysregulation of epigenetic histone modifications

The level of genomic DNA methylation plays an important role in development and disease. In order to establish an experimental system for the functional analysis of genome-wide hypermethylation, we overexpressed the mouse de novo methyltransferase Dnmt3a in Drosophila melanogaster. These flies showed severe developmental defects that could be linked to reduced rates of cell cycle progression and irregular chromosome condensation. In addition, hypermethylated chromosomes revealed elevated rates of histone H3-K9 methylation and a more restricted pattern of H3-S10 phosphorylation. The developmental and chromosomal defects induced by DNA hypermethylation could be rescued by mutant alleles of the histone H3-K9 methyltransferase gene Su(var)3-9. This mutation also resulted in a significantly decreased level of genomic DNA methylation. Our results thus uncover the molecular consequences of genomic hypermethylation and demonstrate a mutual interaction between DNA methylation and histone methylation.     

Capillary electrophoretic analysis of genomic DNA methylation levels

Changes in DNA methylation have been found in the large majority of tumors. This phenomenon includes both genome-wide hypomethylation and gene- specific hypermethylation. However, the clinical relevance of either mechanism has remained contentious. In order to determine DNA methylation levels from a large number of clinical samples, we have established a method for accurate high-throughput quantification of 5-methylcytosine in genomic DNA. Our protocol requires a small amount (<1 µg) of DNA that is enzymatically hydrolyzed to single nucleotides. Single nucleotides are then derivatized with a fluorescent marker and separated by capillary electrophoresis. After calibration of the method, we have determined cytosine methylation levels from tumor samples of 81 patients that had been diagnosed with chronic lymphocytic leukemia (CLL). These patients showed a high variability in their methylation levels with a general trend towards hypomethylation. Because of its high accuracy and throughput our method will be useful in determining the role of genomic DNA methylation levels in tumorigenesis.     

Minireview: On the homology of the protocoel in Cephalochordata and 'lower' Deuterostomia

Hypotheses regarding the homology of the protocoel in planktonic deuterostome larvae and mesodermal structures in ontogenetic stages of cephalochordates are evaluated. The prevalent ‘classical’ hypothesis describes the protocoel as being homologous with the diverticula of Hatschek, which, on the left side, develop into the preoral pit, subsequently into Hatschek’s pit and groove (in part). This hypothesis is based mainly on the position of Hatschek’s diverticula anterior to the rest of the mesoderm during their enterocoelic origin. It is shown here that during development the mesodermal segment that develops into Hatschek’s nephridium is the most anterior one prior to formation of Hatschek’s diverticula, and this segment assumes an anteriormost position after differentiation of Hatschek’s diverticula. Additional similarities between this segment and protocoels are: (i) presence of endomesodermal cells with podocytic extensions, (ii) excretory function, (iii) relatively early ontogenetic origin, (iv) probable lack of association with nervous structures, (v) probable ectodermal origin of a portion of the canal, and (vi) position relative to the mouth opening. Therefore, homology between the protocoel and the segment that becomes Hatschek’s nephridium is proposed. It is concluded that a glandular structure homologous to the diverticula of Hatschek and anterior to the protocoel/Hatschek’s nephridium is a synapomorphy of notochordates or chordates.     

Innervation of the mucosa of the small intestine of laboratory animals. I. Architecture, light microscopic structure and histochemical differentiation

1. The small intestine mucosa has a dense and differentiated innervation. The most dense nerve plexuses are contained in the villi. Each tubular gland is surrounded by a rope-ladder-like plexus, mostly developed in the lower and upper third. 2. According to our observations most of the axons in the mucosa are cholinergic. Cholinergic and adrenergic axons take part in the innervation of the glands. Moreover enterochromaffine cells can be innervated for instance sympathetically. Intraepithelial axons could not be found.     

Interaction of [125I]β nerve growth factor with acidic proteins

We have demonstrated that the nerve growth factor will interact with various acidic proteins apparently nonspecifically. When¹²⁵I-labeled nerve growth factor at a concentration of 3.8×10^–10 M is incubated with an acidic protein at 2 mg/ml (4.5×10^–6–4.4×10^–5 M), a complex is formed. This complex changes the isoelectric point of the¹²⁵I-labeled nerve growth factor sufficiently so that the¹²⁵I-labeled nerve growth factor migrates anomalously in polyacrylamide gel electrophoresis. The interaction between nerve growth factor and bovine serum albumin, which appears to be complex, may be the cause of the previously reported activation of the nerve growth factor when bovine serum albumin is present in a typical bioassay.A preliminary report of this work was presented at the American Society of Biological Chemists, 71st Annual Meeting, in June 1980.     

Real-time imaging of the dynamics of secretory granules in growth cones

Secretory granules containing a hybrid protein consisting of the regulated secretory protein tissue plasminogen activator and an enhanced form of green fluorescent protein were tracked at high spatial resolution in growth cones of differentiated PC12 cells. Tracking shows that granules, unlike synaptic vesicles, generally are mobile in growth cones. Quantitative analysis of trajectories generated by granules revealed two dominant modes of motion: diffusive and directed. Diffusive motion was observed primarily in central and peripheral parts of growth cones, where most granules diffused two to four orders of magnitude more slowly than comparably sized spheres in dilute solution. Directed motion was observed primarily in proximal parts of growth cones, where a subset of granules underwent rapid, directed motion at average speeds comparable to those observed for granules in neurites. This high-resolution view of the dynamics of secretory granules in growth cones provides insight into granule organization and release at nerve terminals. In particular, the mobility of granules suggests that granules, unlike synaptic vesicles, are not tethered stably to cytoskeletal structures in nerve terminals. Moreover, the slow diffusive nature of this mobility suggests that secretory responses involving centrally distributed granules in growth cones will occur slowly, on a time scale of minutes or longer.     

A limited survey of aflatoxins and fumonisins in retail maizebased products in the UK using immunoassay detection

A total of 27 maize-based products destined for human consumption were collected from retail outlets within the city of Glasgow in the UK and were analysed for the presence of aflatoxins using immunoaftinity column chromatography with fluorescence detection and for fumonisins by competitive ELISA. Aflatoxins were detected at a trace level below 4 in eight (30%) of the 27 samples tested, no sample contained aflatoxins at a high level although one sample of sweetcorn did contain aflatoxins at a level of 5-10 Fumonisins were detected in eight (30%) of the samples at levels from 1 to 8mgkg^-1 and a further eight samples contained fumonisin at a level below 1 mgkg^-1 but above the detectable level. The highest concentration of fumonisins was found in a sample of fine corn meal at 8-12mgkg^-1.     

Verrucosispora fiedleri sp. nov., an actinomycete isolated from a fjord sediment which synthesizes proximicins

A novel filamentous actinobacterial organism, designated strain MG-37^T, was isolated from a Norwegian fjord sediment and examined using a polyphasic taxonomic approach. The organism was determined to have chemotaxonomic and morphological properties consistent with its classification in the genus Verrucosispora and formed a distinct phyletic line in the Verrucosispora 16S rRNA gene tree. It was most closely related to Verrucosispora maris DSM 45365^T (99.5 % 16S rRNA gene similarity) and Verrucosispora gifhornensis DSM 44337^T (99.4 % 16S rRNA gene similarity) but was distinguished from these strains based on low levels of DNA:DNA relatedness (~56 and ~50 %, respectively). It was readily delineated from all of the type strains of Verrucosispora species based on a combination of phenotypic properties. Isolate MG-37^T (=NCIMB 14794^T = NRRL-B-24892^T) should therefore be classified as the type strain of a novel species of Verrucosispora for which the name Verrucosispora fiedleri is proposed.     

Genome sequence of the abyssomicin- and proximicin-producing marine actinomycete Verrucosispora maris AB-18-032

Verrucosispora maris AB-18-032 is a marine actinomycete that produces atrop-abyssomicin C and proximicin A, both of which have novel structures and modes of action. In order to understand the biosynthesis of these compounds, to identify further biosynthetic potential, and to facilitate rational improvement of secondary metabolite titers, we have sequenced the complete 6.7-Mb genome of Verrucosispora maris AB-18-032.