Evolution of the Larval Cuticle Proteins Coded by the Secondary Sex Chromosome Pair: X2 and Neo-Y of Drosophila miranda: I. Comparison at the DNA Sequence Level

The larval cuticle protein genes (Lcps) represent a multigene family located at the right arm of the metacentric autosome 2 (2R) in Drosophila melanogaster. Due to a chromosome fusion the Lcp locus of Drosophila miranda is situated on a pair of secondary sex chromosomes, the X2 and neo-Y chromosome. Comparing the DNA sequences from D. miranda and D. melanogaster organization and the gene arrangement of Lcp1–Lcp4 are similar, although the intergene distances vary considerably. The greatest difference between Lcp1 and Lcp2 is due to the occurrence of a pseudogene in D. melanogaster which is not present in D. miranda. Thus the cluster of the four Lcp genes existed already before the separation of the melanogaster and obscura group. Intraspecific homogenizations of different cluster units must have occurred repeatedly between the Lcp1/Lcp2 and Lcp3/Lcp4 sequence types. The most obvious example is exon 2 of the Lcp3 gene in D. miranda, which has been substituted by the corresponding section of the Lcp4 gene rather recently. The homogenization must have occurred before the translocation which generated the neo-Y chromosome. Lcp3 of D. melanogaster has therefore no orthologous partner in D. miranda. Rearrangements in the promoter regions of the D. miranda Lcp genes have generated new, potentially functional CAAT-box motifs. Since three of the Lcp alleles on the neo-Y are not expressed and Lcp3 is expressed only at a reduced level, it is suggestive to speculate that the rearrangements might be involved as cis-regulatory elements in the up-regulation of the X2-chromosomal Lcp alleles, in Drosophila an essential process for dosage compensation. The Lcp genes on the neo-Y chromosome have accumulated more base substitutions than the corresponding alleles on the X2. Received: 27 December 1995 / Accepted: 30 April 1996     

Role of precursor translocation in coordination of murein and phospholipid synthesis in Escherichia coli.

Inhibition of phospholipid synthesis in Escherichia coli by either cerulenin treatment or glycerol starvation of a glycerol-auxotrophic mutant resulted in a concomitant block of murein synthesis. The intracellular pool of cytoplasmic and lipid-linked murein precursors was not affected by an inhibition of phospholipid synthesis, nor was the activity of the penicillin-binding proteins. In addition, a decrease in the activity of the two lipoprotein murein hydrolases, the lytic transglycosylases A and B, could not be demonstrated. The indirect inhibition of murein synthesis by cerulenin resulted in a 68% decrease of trimeric muropeptide structures, proposed to represent the attachment points of newly added murein. Importantly, inhibition of phospholipid synthesis also inhibited O-antigen synthesis with a sensitivity and kinetics similar to those of murein synthesis. It is concluded that the step common for murein and O-antigen synthesis, the translocation of the respective bactoprenolphosphate-linked precursor molecules, is affected by an inhibition of phospholipid synthesis. Consistent with this assumption, it was shown that murein synthesis no longer depends on ongoing phospholipid synthesis in ether-permeabilized cells. We propose that the assembly of a murein-synthesizing machinery, a multienzyme complex consisting of murein hydrolases and synthases, at specific sites of the membrane, where integral membrane proteins such as RodA and FtsW facilitate the translocation of the lipid-linked murein precursors to the periplasm, depends on ongoing phospholipid synthesis. This would explain the well-known phenomenon that both murein synthesis and antibiotic-induced autolysis depend on phospholipid synthesis and thereby indirectly on the stringent control.     

The unexpected long period of elevated CH4 emissions from an inundated fen meadow ended only with the occurrence of cattail (Typha latifolia)

Drainage and agricultural use transform natural peatlands from a net carbon (C) sink to a net C source. Rewetting of peatlands, despite of high methane (CH₄) emissions, holds the potential to mitigate climate change by greatly reducing CO₂ emissions. However, the time span for this transition is unknown because most studies are limited to a few years. Especially, nonpermanent open water areas often created after rewetting, are highly productive. Here, we present 14 consecutive years of CH₄ flux measurements following rewetting of a formerly long-term drained peatland in the Peene valley. Measurements were made at two rewetted sites (non-inundated vs. inundated) using manual chambers. During the study period, significant differences in measured CH₄ emissions occurred. In general, these differences overlapped with stages of ecosystem transition from a cultivated grassland to a polytrophic lake dominated by emergent helophytes, but could also be additionally explained by other variables. This transition started with a rapid vegetation shift from dying cultivated grasses to open water floating and submerged hydrophytes and significantly increased CH₄ emissions. Since 2008, helophytes have gradually spread from the shoreline into the open water area, especially in drier years. This process was periodically delayed by exceptional inundation and eventually resulted in the inundated site being covered by emergent helophytes. While the period between 2009 and 2015 showed exceptionally high CH₄ emissions, these decreased significantly after cattail and other emergent helophytes became dominant at the inundated site. Therefore, CH₄ emissions declined only after 10 years of transition following rewetting, potentially reaching a new steady state. Overall, this study highlights the importance of an integrative approach to understand the shallow lakes CH₄ biogeochemistry, encompassing the entire area with its mosaic of different vegetation forms. This should be ideally done through a study design including proper measurement site allocation as well as long-term measurements.     

Evaluation of Preparation, Staining and Microscopic Techniques for Counting Incremental Lines in Cementum of Human Teeth

Apposition of cementum occurs in phases resulting in two types of layers with different optical and staining properties that can be observed by light microscopy. Narrow, dark staining incremental lines are separated by wider bands of pale staining cementum. The distance from one line to the next represents a yearly increment deposit of cementum in many mammals, and counting these lines has been used routinely to estimate the age of the animals. Incremental lines in cementum have also been observed in sections of human teeth, and the object of the present investigation was to examine a number of methods for preparing and staining them for counting. Longitudinal and transverse sections, either ground or decalcified, were cut from formalin fixed human dental roots, paraffin embedded or frozen, and stained using several techniques. The cementum was investigated using conventional light, fluorescence, polarized light, confocal laser scanning, interference contrast, phase contrast, and scanning electron microscopy. Incremental lines in the cementum could be observed in ground sections and, following decalcification, in both frozen and paraffin embedded sections. Toluidine blue, cresyl violet, hematoxylin, or periodic acid Schiff (PAS) stained incremental lines allowing differentiation by conventional light microscopy. Contrast was best using fluorescence microscopy and excitation by green light since the stained cemental bands, but not the incremental lines, fluoresced after staining with cresyl violet, PAS or hematoxylin and eosin. The results with other microscopic techniques were unsatisfactory. Since incremental lines are not destroyed by acids and stain differently than the remaining cementum, it is likely that they possess an organic structure which differs from the cementum. Incremental lines in human dental cementum could be observed best using decalcified sections stained with cresyl violet excited by green light.     

Influence of central command and ergoreceptors on the splanchnic circulation during isometric exercise

The splanchnic circulation can make a major contribution to blood flow changes. However, the role of the splanchnic circulation in the reflex adjustments to the blood pressure increase during isometric exercise is not well documented. The central command and the muscle chemoreflex are the two major mechanisms involved in the blood pressure response to isometric exercise. This study aimed to examine the behaviour of the superior mesenteric artery during isometric handgrip (IHG) at 30% maximal voluntary contraction (MVC). The pulsatility index (PI) of the blood velocity waveform of the superior mesenteric artery was taken as the study parameter. A total of ten healthy subjects [mean age, 21.1 (SEM 0.3) years] performed an IHG at 30% MVC for 90 s. At 5 s prior to the end of the exercise, muscle circulation was arrested for 90 s to study the effect of the muscle chemoreflex (post exercise arterial occlusion, PEAO). The IHG at 30% MVC caused a decrease in superior mesenteric artery PI, from 4.84 (SEM 1.57) at control level to 3.90 (SEM 1.07) (P = 0.015). The PI further decreased to 3.17 (SEM 0.70) (P = 0.01) during PEAO. Our results indicated that ergoreceptors may be involved in the superior mesenteric artery vasodilatation during isometric exercise.     

Two recurrent nonsense mutations and a 4 bp deletion in a quasi-symmetric element in exon 37 of the NF1 gene

We screened a total of 92 unrelated patients with neurofibromatosis type 1 (NF1) for mutations in exon 37 of the NF1 gene, by using temperature gradient gel electrophoresis. Two novel mutations were found: a 4 bp deletion in a so-called quasi-symmetric element (6789delTTAC) and a recurrent nonsense mutation, which was identified in two unrelated patients, at codon 2264 (C6792A). The independent origin of the latter mutation in two families was confirmed by haplotype analysis. The nonsense mutation and the 4 bp deletion are both predicted to lead to a truncated protein product lacking the Cterminal 20% (aproximately) of its sequence. The occurrence of three independent mutations among 92 NF1 patients at codons 2263–2264 (exon 37) suggests that a specific search for mutations in this area should be undertaken in screening programs for NF1 mutations.     

Circular and linear structures in chromatin diminution of Cyclops

In confirmation of earlier findings, surface-spread early diminution stages of Cyclops furcifer and C. divulsus yield numerous chromatin rings formed by the 25-to 30-nm type of fiber. Their contour lengths have a range of 0.6 16 m in C. divulsus and 0.4–40 m in C. furcifer. Employing the Miller spreading technique nucleosomal chromatin rings were detected in the critical stages of diminution in a size range of 0.6–100 m, though in lower frequencies. Instead, linear fragments of nucleosomal chromatin were found in numbers equal to or surpassing that of the rings.     

The diminution of heterochromatic chromosomal segments in Cyclops (Crustacea,Copepoda)

The chromosomes of Cyclops divulsus, C. furcifer, and C. strenuus, like those of several other Copepods, undergo a striking diminution of chromatin early in embryogenesis. The process is restricted to the presumptive soma cells and occurs at the 5th cleavage in C. divulsus, at the 6th and 7th in C. furcifer, and at the 4th in C. strenuus. The eliminated chromatin derives from the excision of heterochromatic chromosome segments (H-segments). Their chromosomal location is different in the three investigated species: Whereas in C. divulsus and C. furcifer the H-segments form large blocks — exclusively terminal in the former and terminal as well as kinetochoric in the latter — the germ line heterochromatin in C. strenuus is scattered all along the chromosomes. Extensive polymorphism exists with respect to the length of the terminal H-segments in C. furcifer, and with respect to the overall content of heterochromatin in the chromosomes of C. strenuus. In a local race of C. strenuus an extreme form of dimorphism has been found which is sex limited: females as a rule are heterozygous for an entire set of large (heterochromatin-rich), and a second set of small chromosomes in their germ line. Males are homozygous for the large set. In the first three cleavage divisions the H-polymorphism is solely expressed through differences of chromosome length. Following diminution the differences between homologous have disappeared. Feulgen cytophotometry demonstrates that in the three species the 1C DNA value for the germ line, as measured in sperm, is about twice that measured in somatic mitoses (germ line/soma C-values in picograms of DNA: C. strenuus 2.2/0.9, C. furcifer 2.9/1.44, C. divulsus 3.1/1.8). — The data imply that chromatin diminution is based on a mechanism which allows specific DNA segments, regardless of their location and size, to be cut out from the chromosomes without affecting the structural continuity of the remaining DNA. This mechanism may be analogous to that of prokaryotic DNA excision.