Characterization of chicken gizzard calcyclin and examination of its interaction with caldesmon

Using a procedure developed to purify calcyclin from mouse Ehrlich ascites tumor cells calcyclin was purified from smooth muscle of chicken gizzard. Chicken gizzard calcyclin bound to phenyl-Sepharose in a calcium dependent manner as did mouse EAT cells and rabbit lung calcyclin but appeared to be more acidic than its mammalian counterparts as revealed by ion exchange chromatography on Mono Q. Chicken gizzard calcyclin bound ⁴⁵Ca²⁺ on nitrocellulose filters and exhibited a shift in electrophoretic mobility on urea-PAGE depending on Ca²⁺ concentration. Crosslinking experiments with BS³ showed that chicken gizzard calcyclin was able to form noncovalent dimers. As indicated by a decrease in maximum tryptophan fluorescence emission of caldesmon (about 14% at 1:1 molar ratio) and displacement of calmodulin from its complex with caldesmon, chicken gizzard calcyclin binds caldesmon. This binding was, however, much weaker than that of calmodulin and could not influence the interaction of caldesmon with actin. In consequence, calcyclin was unable to reverse the inhibitory effect of caldesmon on actin-activated Mg²⁺-ATPase activity of myosin in the presence of Ca²⁺.     

Analysis of genetic relationships and antimicrobial susceptibility of Escherichia coli isolated from Clethrionomys glareolus

Eleven strains of Escherichia coli were isolated from 54 bank voles living in the LomZa Landscape Park of the Narew River Valley, indicating that E. coli is not common in the alimentary tract of these mammals. On the basis of pulsed-field gel electrophoresis and computer-assisted analysis, the isolates were grouped into six genotypes at similarities of 39%. Chromosome length of E. coli under study differed by as much as 900 kb, ranging 2.7-3.6 Mb. All strains were susceptible to amikacin and ciprofloxacin, whereas, for tetracycline, streptomycin, ampicillin, and cefonicid, different results were noted. No differences were detected among the plasmid complements of eight strains (73%), for which plasmid profiles revealed the presence of two plasmidic bands. One, three and four plasmids were observed in a plasmid pattern of single isolates. The observation from the study indicated the high genetic polymorphism among the isolates recovered from the animals of one species living in the same environment.     

Subspecies of the Eeuropean beaver<Emphasis Type="Italic">Castor fiber</Emphasis> Linnaeus, 1758

Subspecific relationships of the European beaverCastor fiber have been obscured by failure to follow the rules of the International Code of Zoological Nomenclature and remain an open issue. Inconsequence in the use of subspecies names caused a nomenclatural as well as a taxonomic confusion. We discuss these controversial and incorrect nomenclatural decisions, and recognize 9 nominal subspecies of European beaver, bearing 9 available names:C. f. fiber Linnaeus, 1758,C. f. galliae Geoffroy, 1803, C. f. albicus Matschie, 1907,C. f. vistulanus Matschie, 1907,C. f. pohlei Serebrennikov, 1929,C. f. birulai Serebrennikov, 1929,C. f. tuvinicus Lavrov, 1969,C. f. belorussicus Lavrov, 1981, andC. f. orientoeuropaeus Lavrov, 1981.     

Flow Cytometry,HPLC-RP,and metAFLP Analyses to Assess Genetic Variability in Somatic Embryo-Derived Plantlets of <Emphasis Type="BoldItalic">Gentiana pannonica</Emphasis> Scop

Cytometric and molecular techniques were used to verify genetic uniformity among somatic embryo-derived plantlets of Gentiana pannonica Scop. Cytometric analysis of regenerants revealed absence of chromosomal changes and alterations in ploidy. However, reverse phase high pressure liquid chromatography detected higher levels of methylation in regenerated plants than those of control plants. These changes were further investigated using a quantitative molecular marker-based approach. This revealed that numerous tissue culture-induced variations, ∼3% (epi)mutations, were observed, including sequence variation and changes in methylation patterns. Moreover, complex patterns of variation, including combinations of genetic and epigenetic changes, were relatively high (ca. 9%). Overall, tissue culture-induced variation reached 16%; while, demethylation was lower than de novo methylation in heterozygotic material and similar in all regenerated plantlets.     

Ubiquitin hydrolase Uch-L1 rescues beta-amyloid-induced decreases in synaptic function and contextual memory

The neuronal ubiquitin/proteasomal pathway has been implicated in the pathogenesis of Alzheimer's disease (AD). We now show that a component of the pathway, ubiquitin C-terminal hydrolase L1 (Uch-L1), is required for normal synaptic and cognitive function. Transduction of Uch-L1 protein fused to the transduction domain of HIV-transactivator protein (TAT) restores normal enzymatic activity and synaptic function both in hippocampal slices treated with oligomeric Abeta and in the APP/PS1 mouse model of AD. Moreover, intraperitoneal injections with the fusion protein improve the retention of contextual learning in APP/PS1 mice over time. The beneficial effect of the Uch-L1 fusion protein is associated with restoration of normal levels of the PKA-regulatory subunit IIalpha, PKA activity, and CREB phosphorylation.     

Cytogenetic effect of prolonged in vitro exposure of <Emphasis Type="Italic">Allium cepa</Emphasis> L. root meristem cells to salt stress

Salt stress severely affects plant growth and regulation; however knowledge about its effect on cytological changes is limited. In this report, studies were carried out to reveal the morphological and cytogenetic responses of the Allium cepa roots after prolonged (7, 14, 20 days) exposure to salt stress in tissue culture conditions. Roots of plants were treated with different concentrations (100, 200, 300 mM) of NaCl and KCl added to the culture medium. Both salts caused root growth reduction and showed cytotoxic effect reflected in reduction of root tip cells mitotic activity and increase of different abnormalities such as chromosome breaks and bridges and formation of micronuclei. This experiment showed differences of the action of excessive amounts of both tested salts at the cellular level, as KCl induced higher frequencies of abnormalities during cell divisions, whereas NaCl showed more mitodepressive effect and more frequently led to the root meristem cell death.     

Neonatal susceptibility to UV induced cutaneous malignant melanoma in a mouse model.

UV irradiation has multiple effects on skin including erythema, immunosuppression and the induction of keratinocyte-derived skin cancers and cutaneous malignant melanoma (CMM). CMM which arises from damage to the melanocyte, the pigment cell of the skin, is associated in epidemiologic studies with sun-exposure of susceptible populations, especially children. Our experimental studies have supported the concept that the epidemiologically observed susceptibility in children has a biologic basis. Hepatocyte growth factor/scatter factor (HGF/SF) transgenic mice neonatally irradiated with UV produce melanomas which recapitulate human disease in histopathology and molecular pathogenesis. In this model, neonatal UV is necessary and sufficient for melanoma induction although an additional adult dose of UV radiation significantly increased melanoma multiplicity. One hypothesis for the susceptibility of neonatal mice to induction of melanoma is that neonatal skin contains a large number of immature melanocytes which may result in the retention of the consequences of UV damage throughout the lifetime of the animal. An alternate hypothesis is that the immaturity of the neonatal immune system results in tolerance to melanocytic antigens produced by UV exposure, thus permitting the subsequent outgrowth of melanoma. Here, we discuss the current state of knowledge about the differences between adult and neonatal mice in melanocytes and immune maturation as possible factors playing a role in the susceptibility to melanoma in UV irradiated HGF/SF transgenic mice.     

Topological model of the division process of cellular and subcellular compartments

The application of the theory of homeomorphic transformations of topological manifolds and the operation of the connected sum of manifolds for a formation of a topological model of membrane transformations during the division process of cellular and subcellular compartments, has been shown. The biological cell and the subcellular structures in the form of vesicles are modelled by an arrangement of two concentric spheres corresponding to the inner and outer layer of the membrane bounding the vesicle. The analysis shows eight succeeding topological stages of membrane transformations during the division process and these stages are characterised. It is concluded that there is a vectorial translocation of lipid molecules from the inner layer of the membrane bounding the vesicle before the division process to the outer layer of the membranes after the division process and there is no lipid translocation from the outer layer to the inner layers during the division process.     

Synthesis of gamma- and delta-lactones from 1alpha-hydroxy-5,6-trans-vitamin D3 by ring-closing metathesis route and their reduction with metal hydrides

New synthetic pathway towards 19-functionalized derivatives of 1alpha-hydroxy-5,6-trans-vitamin D3 was described. Ring-closing metathesis (RCM) of 1alpha-hydroxy-5,6-trans-vitamin D3 1-omega-alkenoates was a key-step. Hydride reduction of resulting lactones led to the new vitamin D3 analogues.     

Conventional kinesin holoenzymes are composed of heavy and light chain homodimers

Conventional kinesin is a major microtubule-based motor protein responsible for anterograde transport of various membrane-bounded organelles (MBO) along axons. Structurally, this molecular motor protein is a tetrameric complex composed of two heavy (kinesin-1) chains and two light chain (KLC) subunits. The products of three kinesin-1 (kinesin-1A, -1B, and -1C, formerly KIF5A, -B, and -C) and two KLC (KLC1, KLC2) genes are expressed in mammalian nervous tissue, but the functional significance of this subunit heterogeneity remains unknown. In this work, we examine all possible combinations among conventional kinesin subunits in brain tissue. In sharp contrast with previous reports, immunoprecipitation experiments here demonstrate that conventional kinesin holoenzymes are formed of kinesin-1 homodimers. Similar experiments confirmed previous findings of KLC homodimerization. Additionally, no specificity was found in the interaction between kinesin-1s and KLCs, suggesting the existence of six variant forms of conventional kinesin, as defined by their gene product composition. Subcellular fractionation studies indicate that such variants associate with biochemically different MBOs and further suggest a role of kinesin-1s in the targeting of conventional kinesin holoenzymes to specific MBO cargoes. Taken together, our data address the combination of subunits that characterize endogenous conventional kinesin. Findings on the composition and subunit organization of conventional kinesin as described here provide a molecular basis for the regulation of axonal transport and delivery of selected MBOs to discrete subcellular locations.