Allozyme polymorphism of Mdh and α-Gpdh in Ixodes ricinus populations: comparison of borreliae-infected and uninfected ticks

Ixodes ricinus Linnaeus (Acari: Ixodidae) ticks are vectors of numerous infectious diseases in humans and animals. The allozyme variability of MDH and α-Gpdh was detected by native polyacrylamide gel electrophoresis in I. ricinus natural populations in three localities in Serbia. Four alleles of Mdh locus (MDH 1, MDH 2, MDH 3 and MDH X) and four alleles of α-Gpdh locus (VS, S, F and VF) were detected. Interpopulation differences in Mdh and α-Gpdh allele frequencies were statistically insignificant. Significant difference in α-Gpdh allele frequencies between males and females was recorded in the largest sample only. Differences in allele frequencies, detected between borreliae-infected and uninfected I. ricinus ticks, were close to the level of statistical significance, especially for α-Gpdh locus. Clear significant difference appeared in females when sexes were tested separatelly (P = 0.037). It is interesting that genotypes containing rarer alleles (MDH 1 and S) were infected in higher proportion in comparison to other genotypes. Our results point towards a possible role of Mdh and α-Gpdh loci in I. ricinus ticks in the determination of energy requirements for host seeking. Sex differences in α-Gpdh allele frequencies suggest that selective pressure, concerning efficiency of reserve materials utilisation, points to α-Gpdh rather than to Mdh locus.     

Reappearance of catatonia. A case report

We describe a patient who was ill for more than 15 years. She was treated predominantly as a neurological case because of multiple motor signs. The diagnosis of catatonia was considered at the time when she was hospitalized in a psychiatric hospital after a suicidal attempt. The "therapeutic blindness", which was obviously present during 15 years of her illness, is discussed: the ICD--10 is perhaps misleading regarding this diagnosis while it associates it too tightly only to schizophrenia.     

Asexual Reproduction and Segmental Regeneration, but not Morphallaxis, are Inhibited by Boric Acid in Lumbriculus variegatus (Annelida: Clitellata: Lumbriculidae)

Body fragmentation, in some animal groups, is a mechanism for survival and asexual reproduction. Lumbriculus variegatus (Müller, 1774), an aquatic oligochaete worm, is capable of regenerating into complete individuals from small body fragments following injury and reproduces primarily by asexual reproduction. Few studies have determined the cellular mechanisms that underlie fragmentation, either regenerative or asexual. We utilized boric acid treatment, which blocks regeneration of segments in amputated fragments and blocks architomic fission during asexual reproduction, to investigate mechanistic relationships and differences between these two modes of development. Neural morphallaxis, involving changes in sensory fields and giant fiber conduction, was detected in amputated fragments in the absence of segmental regeneration. Furthermore, neural morphallactic changes occurred as a result of developmental mechanisms of asexual reproduction, even when architomy was prevented. These results show that fragmentation in L. variegatus, during injury or asexual reproduction, employs developmental and morphallactic processes that can be mechanistically dissociated by boric acid exposure. In regeneration following injury, compensatory morphallaxis occurred in response to fragmentation. In contrast, anticipatory morphallaxis was induced in preparation for fragmentation during asexual reproduction. Thus, various forms of regeneration in this lumbriculid worm can be activated independently and in different developmental contexts.     

A mutation linked with autism reveals a common mechanism of endoplasmic reticulum retention for the alpha,beta-hydrolase fold protein family

A mutation linked to autistic spectrum disorders encodes an Arg to Cys replacement in the C-terminal portion of the extracellular domain of neuroligin-3. The solvent-exposed Cys causes virtually complete retention of the protein in the endoplasmic reticulum when the protein is expressed in transfected cells. An identical Cys substitution was reported for butyrylcholinesterase through genotyping patients with post-succinylcholine apnea. Neuroligin, butyrylcholinesterase, and acetylcholinesterase are members of the alpha,beta-hydrolase fold family of proteins sharing sequence similarity and common tertiary structures. Although these proteins have distinct oligomeric assemblies and cellular dispositions, homologous Arg residues in neuroligin-3 (Arg-451), in butyrylcholinesterase (Arg-386), and in acetylcholinesterase (Arg-395) are conserved in all studied mammalian species. To examine whether an homologous Arg to Cys mutation affects related proteins similarly despite their differing capacities to oligomerize, we inserted homologous mutations in the acetylcholinesterase and butyrylcholinesterase cDNAs. Using confocal fluorescence microscopy and analysis of oligosaccharide processing, we find that the homologous Arg to Cys mutation also results in endoplasmic reticulum retention of the two cholinesterases. Small quantities of mutated acetylcholinesterase exported from the cell retain activity but show a greater K(m), a much smaller k(cat), and altered substrate inhibition. The nascent proteins associate with chaperones during processing, but the mutation presumably restricts processing through the endoplasmic reticulum and Golgi apparatus, because of local protein misfolding and inability to oligomerize. The mutation may alter the capacity of these proteins to dissociate from their chaperone prior to oligomerization and processing for export.     

Filamin-a and rheological properties of cultured melanoma cells

Here we report the rheological properties of cultured hsFLNa (filamin-A)-expressing (FIL+) and hsFLNa-deficient (FIL-) melanoma cells. Using magnetic twisting cytometry over a wide range of probing frequencies, and targeting either cortical or deeper cytoskeletal structures, we found that differences in stiffness of FIL+ versus FIL- cells were remarkably small. When probed through deep cytoskeletal structures, FIL+ cells were, at most, 30% stiffer than FIL- cells, whereas when probed through more peripheral cytoskeletal structures FIL- cells were not different except at very high frequencies. The loss tangent, expressed as an effective cytoskeletal temperature, was systematically greater in FIL- than FIL+ cells, but these differences were small and showed that the FIL+ cells were only slightly closer to a solidlike state. To quantify cytoskeletal remodeling, we measured spontaneous motions of beads bound to cortical cytoskeletal structures and found no difference in FIL+ versus FIL- cells. Although mechanical differences between FIL+ and FIL- cells were evident both in cortical and deeper structures, these differences were far smaller than expected based on measurements of the rheology of purified actin-filamin solutions. These findings do not rule out an important contribution of filamin to the mechanical properties of the cortical cytoskeleton, but suggest that effects of filamin in the cortex are not exerted on the length scale of the probe used here. These findings would appear to rule out any important contribution of filamin to the bulk mechanical properties of the cytoplasm, however. Although filamin is present in the cytoplasm, it may be inactive, its mechanical effects may be small compared with other crosslinkers, or mechanical properties of the matrix may be dominated by an overriding role of cytoskeletal prestress.     

Influence of the temperature regime on the composition of the macrozoobenthos community in a thermal brook in Serbia

In contrast to cold and eurythermal waters, benthic communities of warm brooks in temperate regions have been inadequately studied. In order to investigate the effects of water thermal regime on the benthic communities of warm waters and their relationships with those of cold and eurythermic ones, the macrozoobenthos was studied at eight sites in the Toplica River, and at four sites in its tributary, the Termalni brook. Investigations were carried out seasonally from April 2000 to January 2001. Warm waters of the Termalni brook were characterized by specific macrozoobenthos assemblages that exhibited significant differences to the populations of eurythermal and cold waters of the Toplica River. The dominant taxa in the macrozoobenthos community of warm waters were mainly Gastropoda species. Moreover, benthic communities of warm waters were characterized by lower diversity and greater biomass in comparison with those of cold and eurythermal waters. The gradient of average annual temperatures represented the main ecological factor influencing changes of diversity and biomass along the course of the investigated Termalni brook. Inflow of warm waters at site T₆ lead to a decrease in macrozoobenthos abundance and changes in qualitative and quantitative composition of the benthocoenosis of a highland stream, but did not significantly alter diversity.     

Identification of a novel glycosyltransferase involved in LOS biosynthesis of Moraxella catarrhalis

Moraxella catarrhalis is an important human mucosal pathogen that contributes to otitis media in infants and exacerbates conditions such as chronic obstructive pulmonary disease in the elderly. This study describes the identification of a novel gene, lgt5 that encodes a glycosyltransferase involved in the LOS biosynthesis of M. catarrhalis. Analysis of NMR data of LOS-derived oligosaccharide from a Serotype A lgt5 mutant strain of M. catarrhalis indicate that lgt5 encodes an alpha-(1-->4)-galactosyltransferase.     

Fractures in late medieval skeletal populations from Serbia

Bone fractures were analyzed from skeletal remains of 861 adult individuals from six cemeteries dating to the Late Medieval period in Serbia. Results of the study were compared to other cemetery populations (635 individual skeletons) of the same date and region in an attempt to understand fracture patterns. The association of types of fractures and their prevalence with sex, age at death, cemetery site, and information deriving from historical sources are discussed. Results showed that the long bone fracture frequency was 0.7%, and the majority of the fractures were the result of direct force. This rate is similar to some studies of contemporary British skeletal samples. However, it is much lower than for some other Old World sites. Cranial vault fractures had a rate of 6.7%, and of the facial skeleton, 1.3%; the frontal bone was the most affected of bones of the cranial vault. Injuries were more common on the upper extremities (0.8%) compared to the lower (0.6%). However, the fibula was the most fractured bone (2.8%), followed by the ulna (2.4%). This pattern is similar to three of six Late Medieval urban sites in Britain. These findings suggest that this rural community was exposed to a low risk of trauma, probably related mostly to accidents sustained during farming, and rarely to interpersonal violence.     

Length-dependent prediction of protein intrinsic disorder

Background  

Due to the functional importance of intrinsically disordered proteins or protein regions, prediction of intrinsic protein disorder from amino acid sequence has become an area of active research as witnessed in the 6th experiment on Critical Assessment of Techniques for Protein Structure Prediction (CASP6). Since the initial work by Romero et al. (Identifying disordered regions in proteins from amino acid sequences, IEEE Int. Conf. Neural Netw., 1997), our group has developed several predictors optimized for long disordered regions (>30 residues) with prediction accuracy exceeding 85%. However, these predictors are less successful on short disordered regions (≤30 residues). A probable cause is a length-dependent amino acid compositions and sequence properties of disordered regions.     

Covalent inhibition of hAChE by organophosphates causes homodimer dissociation through long-range allosteric effects

Acetylcholinesterase (EC 3.1.1.7), a key acetylcholine-hydrolyzing enzyme in cholinergic neurotransmission, is present in a variety of states in situ, including monomers, C-terminally disulfide-linked homodimers, homotetramers, and up to three tetramers covalently attached to structural subunits. Could oligomerization that ensures high local concentrations of catalytic sites necessary for efficient neurotransmission be affected by environmental factors? Using small-angle X-ray scattering (SAXS) and cryo-EM, we demonstrate that homodimerization of recombinant monomeric human acetylcholinesterase (hAChE) in solution occurs through a C-terminal four-helix bundle at micromolar concentrations. We show that diethylphosphorylation of the active serine in the catalytic gorge or isopropylmethylphosphonylation by the R_P enantiomer of sarin promotes a 10-fold increase in homodimer dissociation. We also demonstrate the dissociation of organophosphate (OP)-conjugated dimers is reversed by structurally diverse oximes 2PAM, HI6, or RS194B, as demonstrated by SAXS of diethylphosphoryl-hAChE. However, binding of oximes to the native ligand-free hAChE, binding of high-affinity reversible ligands, or formation of an S_P-sarin-hAChE conjugate had no effect on homodimerization. Dissociation monitored by time-resolved SAXS occurs in milliseconds, consistent with rates of hAChE covalent inhibition. OP-induced dissociation was not observed in the SAXS profiles of the double-mutant Y337A/F338A, where the active center gorge volume is larger than in wildtype hAChE. These observations suggest a key role of the tightly packed acyl pocket in allosterically triggered OP-induced dimer dissociation, with the potential for local reduction of acetylcholine-hydrolytic power in situ. Computational models predict allosteric correlated motions extending from the acyl pocket toward the four-helix bundle dimerization interface 25 Å away.