"The orchids have been a splendid sport"--an alternative look at Charles Darwin's contribution to orchid biology

Charles Darwin's work with orchids and his thoughts about them are of great interest and not a little pride for those who are interested in these plants, but they are generally less well known than some of his other studies and ideas. Much has been published on what led to his other books and views. However, there is a paucity of information in the general literature on how Darwin's orchid book came about. This review will describe how The Various Contrivances by Which Orchids Are Fertilised by Insects came into being and will discuss the taxonomy of the orchids he studied. It also will concentrate on some of the less well-known aspects of Darwin's work and observations on orchids-namely, rostellum, seeds and their germination, pollination effects, and resupination-and their influence on subsequent investigators, plant physiology, and orchid science.     

Developmental consequences of imprinting of parental chromosomes by DNA methylation

Genomic imprinting by epigenetic modifications, such as DNA methylation, confers functional differences on parental chromosomes during development so that neither the male nor the female genome is by itself totipotential. We propose that maternal chromosomes are needed at the time when embryonic cells are totipotential or pluripotential, but paternal chromosomes are probably required for the proliferation of progenitor cells of differentiated tissues. Selective elimination or proliferation of embryonic cells may occur if there is an imbalance in the parental origin of some alleles. The inheritance of repressed and derepressed chromatin structures probably constitutes the initial germ-line-dependent 'imprints'. The subsequent modifications, such as changes in DNA methylation during early development, will be affected by the initial inheritance of epigenetic modifications and by the genotype-specific modifier genes. A significant number of transgene inserts are prone to reversible methylation imprinting so that paternally transmitted transgenes are undermethylated, whereas maternal transmission results in hypermethylation. Hence, allelic differences in epigenetic modifications can affect their potential for expression. The germ line evidently reverses the previously acquired epigenetic modifications before the introduction of new modifications. Errors in the reversal process could result in the transmission of epigenetic modifications to subsequent generation(s) with consequent cumulative phenotypic and grandparental effects.     

ABO blood typing of human skeletal remains in Hungary

The author reports about the theoretical effects of his paleoserologic investigations on some historical population genetics problems. First he refers to the essence of the two modifications by the help of which the fluorescent antibody method can be made suitable for blood typing or archeological skeletal remains and determines his working units (sample, series, "population") used in the paleoserologic researches. The benefits of the projection of the ABO blood typing results on the map of the cemetaries are demonstrated. The distribution of the several phenotypes are collated to the character or richness of the grave goods and to the taxonomic features of the late individuals. The thorough examination of the serogenetic distances among the several samples of a given historical period may cast more light on the ethnic interrelations of the earlier populations living in the same geographic area. Following up the serogenetic changes of a population during subsequent historical periods, new ideas can be gained about the importance of the environmental, economic, and demographic factors shaping the serogenetic profile of the population.     

The "concrete" histo-cytological diagnosis of noninfiltrative cancer

The diagnosis of noninfiltrative carcinomas (intraepithelial carcinoma, respectively intramucosal adenocarcinoma) is based on the existence of architectural modifications of the stratified squamous epithelia (the skin included) and of glandular structures. The architectural alterations are associated with cellular, respectively nucleo-cytoplasmic changes. The most useful cellular modifications for the diagnosis of the noninfiltrative malignant process of epithelial origin are: the cellular pleomorphism (anisocytosis and especially anisokaryosis), large nuclei in comparison with cell dimensions determining modifications of the nucleo-cytoplasmic ratio, the presence of hypertrophic nucleoli, respectively giant and/or multiple nucleoli with the change of the nucleolo-nuclear ratio in favour of the nucleolus, a large percentage of mitoses (more than 20 mitoses per thousand cells), respectively more than 4 mitoses in the microscopic field at a high magnification.     

Variations on the topic of the "histone code"

Histones are the fundamental structural proteins intimately associated with eukaryotic DNA to form a highly ordered and condensed nucleoproteic complex termed chromatin. They are the targets of various posttranslational modifications including acetylation, methylation, phosphorylation and ubiquitination that modulate the structure/function of chromatin. The combinatorial nature of histone modifications is hypothesized to define a "histone code" that considerably extends the information potential of the genetic code, giving rise to epigenetic information. Moreover, most core histones consist of several nonallelic variants that can mark specific loci and could play an important role in establishment and maintenance of epigenetic memory. Here we will briefly present our current knowledge about histone posttranslational modifications and their implications in the regulation of epigenetic information. We will next describe core histone variants, insisting on their mode of incorporation into chromatin to discuss their epigenetic function and inheritance.     

Foreleg Autotomy Reduces Mating Success of Male <Emphasis Type="Italic">Schizocosa ocreata</Emphasis> Wolf Spiders

Autotomy is a taxonomically widespread antipredator tactic that allows animals to escape life-threatening situations. Opposing the benefits of survival, animals that have autotomized appendages may later suffer reduced ability in important determinants of fitness. Male Schizocosa ocreata wolf spiders use their forelegs during courtship for visual displays, for tactile courtship, and to defend against attacks by females. In nature they are often found missing one, and sometimes both, forelegs. We found that autotomy of one foreleg has little effect on male ability to mate with virgin females, but that autotomy of both forelegs causes a significant reduction in mating success. Among males that mated, autotomy of one or both forelegs did not influence latency until mating, period spent mounted, probability that his mate would accept a subsequent suitor, or probability that his mate would kill a subsequent suitor.     

A follow-up to "Anti-cytokine therapy in chronic destructive arthritis" by Wim B van den Berg

In recent years, the effectiveness of anti-TNF therapy in treating rheumatoid arthritis (RA) has become apparent. While trials of IL-1 receptor antagonist in RA have been encouraging, it clearly is more difficult to target two molecules (IL-1 α and β) than one (TNF-α). In his review article, Professor Wim van den Berg argues that both TNF-α and IL-1 must be blocked in RA and that although TNF is clearly a potent inflammatory molecule, the dominant cytokine in the subsequent degradation of the joint tissue is IL-1. This commentary discusses his hypothesis in light of animal studies and the limitations of the conclusions that can be drawn from them. More broadly, it discusses the biology of TNF-α and IL-1 and suggests explanations of why TNF-α is a pivotal cytokine in this disease.     

Microsurgical forearm "cricket bat-transformer" phalloplasty.

Presently, the donor flap of choice for microsurgical phallic reconstruction is the radial forearm flap. The success of several different design modifications confirms the reliability of the radial and ulnar forearm flaps. Farrow et al. described their experience with the "cricket bat" concept in 1980. To the previous "cricket bat" design, we now wish to add modifications. These modifications utilize longitudinal and transverse rotations of the linear forearm tissues to create a phallus--much like the transformation of a toy robot into a truck. Deepithelialized flaps and a full-thickness skin graft coronoplasty complete glans reconstruction. The "cricket bat-transformer" flap appears to produce the most predictable results in subtotal phallic reconstructions and phallic constructions in the pediatric and transgender patient groups.     

RNA immunoprecipitation technique for <Emphasis Type="Italic">Drosophila melanogaster</Emphasis> S2 cells

RNA-binding proteins play an important role in RNA metabolism, especially in mRNA biogenesis and subsequent expression patterns regulation. RNA immunoprecipitation (RIP) is a powerful tool for detecting protein–RNA associations. In this paper, we briefly cover the history of this method for analyzing RNA–protein interactions and reviewing a number of modifications of the RIP technique. We also present an adjusted RIP protocol that was modified for Drosophila S2 cell culture. The use of this protocol allows one to perform the efficient precipitation of RNA–protein complexes and harvest RNA in amounts that are sufficient for its downstream analysis.     

Biosynthesis and enzymology of the Caenorhabditis elegans cuticle: identification and characterization of a novel serine protease inhibitor

Caenorhabditis elegans represents an excellent model in which to dissect the biosynthesis and assembly of the nematode cuticle. A sequenced genome, straightforward transgenesis, available mutants and practical genome-wide RNAi approaches provide an invaluable toolkit in the characterization of cuticle components. We have performed a targeted RNAi screen in an attempt to identify components of the cuticle collagen biosynthetic pathway. Collagen biosynthesis and cuticle assembly are multi-step processes that involve numerous key enzymes involved in post-translational modification, trimer folding, procollagen processing and subsequent cross-linking stages. For many of these steps, the modifications and the enzymes are unique to nematodes and may represent attractive targets for the control of parasitic nematodes. A novel serine protease inhibitor was uncovered during our targeted screen, which is involved in collagen maturation, proper cuticle assembly and the moulting process. We have confirmed a link between this inhibitor and the previously uncharacterised bli-5 locus in C. elegans. The mutant phenotype, spatial expression pattern and the over-expression phenotype of the BLI-5 protease inhibitor and their relevance to collagen biosynthesis are discussed.     

"Glyco-deglyco" processes during the biosynthesis of glycoproteins

For the past fifteen years, it has appeared increasingly evident that the N-glycosylation process was accompanied by the release of oligomannoside type oligosaccharides. This material is constituted of oligosaccharide-phosphates and of neutral oligosaccharides possessing one GlcNAc (OS-Gn1) or two GlcNAc (OS-Gn2) at the reducing end. It has been demonstrated that oligosaccharide-phosphates originated from the cleavage, by a specific pyrophosphatase, of non-glucosylated cytosolic faced oligosaccharide-PP-Dol and chiefly the Man5GlcNAc2-PP-Dol. The Man5GlcNAc2-P, as the main product, is recovered in the cytosolic compartment and is further degraded to Man5GlcNAc1 by not-yet depicted enzymes. In contrast, OS-Gn2 produced from hydrolysis of oligosaccharide-PP-Dol (presumably as a transfer reaction onto water) when the amount of protein acceptor is limiting, are generated into the lumen of rough endoplasmic reticulum (rough ER). They are further submitted to processing a-glucosidases and rough ER mannosidase and are (mainly as Man8GlcNAc2) exported into the cytosolic compartment. This material is further degraded into a single component, the Man5GlcNAc1, by the sequential action of a cytosolic neutral chitobiase followed by cytosolic mannosidase(s). Furthermore, OS-Gn1 could have a dual origin: in one hand, they originate from OS-Gn2 by the cytosolic degradation pathway indicated above, on the other hand, we will discuss a possible origin from the degradation or remodelling of newly synthesized glycoproteins. Considered first as a minor phenomenon, these observations have lead to the concept of intracellular oligomannoside trafficking, a process which results from more fundamental phenomena such as the control of the dolichol cycle, and the so-called quality-control of glycoprotein. In this review, we would like to describe the evolution of ideas on the origin, intracellular trafficking and putative roles of these oligomannosides released during during the N-glycosylation process. We propose that these early stage "glyco-deglyco" processes represent a way of control of N-glycosylation and of the fate of N-glycoproteins. This review is dedicated to Pr Paul Boulanger who has spent a large part of his career to determine the structure of proteins in order to understand their functions. If it is well established that many biological functions are born by proteins, it appears more and more evident that co- or post translational modifications are of importance in the modulation of these functions. Among them, the glycosylation appears as a major event which intervene in the 3D structure of the protein, which control his biological time-life and which may act in many recognition processes.     

The "unfrozen fraction" hypothesis of freezing injury to human erythrocytes: a critical examination of the evidence

This paper examines the experimental evidence presented by Mazur and his colleagues to support their hypothesis that the survival of slowly frozen human red blood cells is primarily dependent on the fraction of water that remains unfrozen, rather than on the high concentrations of sodium chloride produced by the formation of ice. This hypothesis is in direct conflict with the general belief that freezing injury under such conditions is caused by the concentration of solutes in the solution surrounding the cells: if the "unfrozen fraction" hypothesis is true, then much of the evidence supporting that belief must be dismissed as mere coincidence. We have reexamined Mazur's data, and have suggested an alternative explanation--that cells which are initially suspended in solutions that are not isotonic differ in their susceptibility to subsequent freezing and thawing, shrunken cells being more resistant and swollen cells more susceptible than normal cells. If this is true then the data can be explained without invoking a direct effect of the unfrozen fraction, solely on the basis of changes in the concentration of the solution surrounding the cells. We cite other experimental evidence, obtained in the absence of freezing, that red blood cells do indeed possess the required property. We further argue that the known effects of variations in cooling and warming rate, and in hematocrit, are able to account for the features observed by Mazur and his colleagues in their three published studies.     

Origins and formation of histone methylation across the human cell cycle

The connections between various nuclear processes and specific histone posttranslational modifications are dependent to a large extent on the acquisition of those modifications after histone synthesis. The reestablishment of histone posttranslational modifications after S phase is especially critical for H3K9 and H3K27 trimethylation, both of which are linked with epigenetic memory and must be stably transmitted from one cellular generation to the next. This report uses a proteomic strategy to interrogate how and when the cell coordinates the formation of histone posttranslational modifications during division. Paramount among the findings is that H3K9 and H3K27 trimethylation begins during S phase but is completed only during the subsequent G(1) phase via two distinct pathways from the unmodified and preexisting dimethylated states. In short, we have systematically characterized the temporal origins and methylation pathways for histone posttranslational modifications during the cell cycle.     

A novel, non-substrate-based series of glycine type 1 transporter inhibitors derived from high-throughput screening

The synthesis and structure-activity relationships (SAR) of a series of indane and tetralin inhibitors of the type 1 glycine transporter, derived from a high-throughput screening (HTS) hit, are described. Key modifications that reduced the 5HT1B receptor affinity of the HTS hit and the P450 2D6 inhibition of subsequent analogues are delineated. While these modifications led to potent and selective GlyT1 inhibitors, HERG affinity and human microsomal clearance remain an issue for this series of compounds.     

Creation of "super" glucocorticoid receptors by point mutations in the steroid binding domain

Almost all modifications of the steroid binding domain of glucocorticoid receptors are known to cause a reduction or loss of steroid binding activity. Nonetheless, we now report that mutations of cysteine 656 of the rat receptor, which was previously suspected to be a crucial amino acid for the binding process, have produced "super" receptors. These receptors displayed an increased affinity for glucocorticoid steroids and a decreased relative affinity for cross-reacting steroids such as progesterone and aldosterone. The increased in vitro affinity of the super receptors was maintained in a whole cell bioassay. These results indicate that additional modifications of the glucocorticoid receptor, and probably the other steroid receptors, may further increase the binding affinity and/or specificity.     

Attempt at ecological prognosis of the plankton in the man-made lake ‘Parana medio (Chapeton transect)’, Argentina

Characteristics of the projected man-made lake on the Middle Parana are described, as well as environmental conditions in the river and their modifications in the future lake. The sources of the future plankton are considered. The formation of the future pelagic ecosystem, mainly of the zooplankton, during the initial and the subsequent stage is discussed.     

Typification of Nordic vascular plants. 1. Names published by G. Wahlenberg

Names of sixtyone taxa described by G. Wahlenberg based on Nordic material are typified. An additional four are listed but no type material is traced. Wahlenberg's four journeys and various modifications of his handwriting are presented. Flora nordica notes no. 4     

Synthesis of phlorizin derivatives and their inhibitory effect on the renal sodium/d-glucose cotransport system

To characterize further the Na⁺/d-glucose cotransport system in renal brush border membranes, phlorizin - a potent inhibitor of d-glucose transport - has been chemically modified without affecting the d-glucose moiety or changing the side groups that are essential for the binding of phlorizin to the Na⁺/d-glucose cotransport system. One series of chemical modifications involved the preparation of 3-nitrophlorizin and the subsequent catalytic reduction of the nitro compound to 3-aminophlorizin. From 3-aminophlorizin, 3-bromoacetamido-, 3-dansyl- and 3-azidophlorizin have been synthesized. In another approach, 3′-mercuryphlorizin was obtained by reaction of phlorizin with Hg(II) acetate. The phlorizin derivatives inhibit sodium-dependent but not sodium-independent d-glucose uptake by hog renal brush border membrane vesicles in the following order of potency: 3′-mercuryphlorizin = phlorizin > 3-aminophlorizin > 3-bromoacetamidophlorizin > 3-azidophlorizin > 3-nitrophlorizin > 3-dansylphlorizin. 3-Bromoacetamidophlorizin - a potential affinity label - also inhibits sodium-dependent but not sodium-independent phlorizin binding to brush border membranes. In addition, sodium-dependent phosphate and sodium-dependent alanine uptake are not affected by 3-bromoacetamidophlorizin. The results described above indicate that specific modifications of the phlorizin molecule at the A-ring or B-ring are possible that yield phlorizin derivatives with a high affinity and high specificity for the renal Na⁺/d-glucose cotransport system. Such compounds should be useful in future studies using affinity labeling (3-bromoacetamido- and 3-azidophlorizin) or fluorescent probes (3-dansylphlorizin).     

Protein modifications during antiviral heat bioprocessing and subsequent storage

Antiviral heat treatment is routinely used in the bioprocessing of therapeutic proteins as a means of reducing viral load. However, in protein formulations containing sucrose this form of bioprocessing can lead to protein modifications. Using a model protein, hen egg white lysozyme, we investigated the effects of antiviral heat treatments in the presence of sucrose on protein integrity during subsequent long-term protein storage. Although heat treatment alone resulted in protein modification, subsequent medium- to long-term storage of both lyophilized and liquid samples at room temperature or above led to further protein modifications. The majority of these modifications were due to the formation of glycation and advanced glycation end products via the reaction of reducing sugars and their autoxidation products (derived from hydrolyzed sucrose) with function groups on the protein surface. These findings have implications for the improvement of therapeutic protein bioprocessing to ensure protein product quality.