Characterization of a new mouse model for peripheral T cell lymphoma in humans

Peripheral T cell lymphomas (PTCLs) are associated with a poor prognosis due to often advanced disease at the time of diagnosis and due to a lack of efficient therapeutic options. Therefore, appropriate animal models of PTCL are vital to improve clinical management of this disease. Here, we describe a monoclonal CD8(+) CD4(-) αβ T cell receptor Vβ2(+) CD28(+) T cell lymphoma line, termed T8-28. T8-28 cells were isolated from an un-manipulated adult BALB/c mouse housed under standard pathogen-free conditions. T8-28 cells induced terminal malignancy upon adoptive transfer into syngeneic BALB/c mice. Despite intracellular expression of the cytotoxic T cell differentiation marker granzyme B, T8-28 cells appeared to be defective with respect to cytotoxic activity as read-out in vitro. Among the protocols tested, only addition of interleukin 2 in vitro could partially compensate for the in vivo micro-milieu in promoting growth of the T8-28 lymphoma cells.     

Experimental determination of the vertical alignment between the second and third transmembrane segments of muscle nicotinic acetylcholine receptors

Nicotinic acetylcholine receptors (nAChR) are members of the Cys‐loop ligand‐gated ion channel superfamily. Muscle nAChR are heteropentamers that assemble from two α, and one each of β, γ, and δ subunits. Each subunit is composed of three domains, extracellular, transmembrane and intracellular. The transmembrane domain consists of four α‐helical segments (M1–M4). Pioneering structural information was obtained using electronmicroscopy of Torpedo nAChR. The recently solved X‐ray structure of the first eukaryotic Cys‐loop receptor, a truncated (intracellular domain missing) glutamate‐gated chloride channel α (GluClα) showed the same overall architecture. However, a significant difference with regard to the vertical alignment between the channel‐lining segment M2 and segment M3 was observed. Here, we used functional studies utilizing disulfide trapping experiments in muscle nAChR to determine the spatial orientation between M2 and M3. Our results are in agreement with the vertical alignment as obtained when using the GluClα structure as a template to homology model muscle nAChR, however, they cannot be reconciled with the current Torpedo nAChR model. The vertical M2–M3 alignments as observed in X‐ray structures of prokaryotic Gloeobacter violaceus ligand‐gated ion channel and GluClα are in agreement. Our results further confirm that this alignment in Cys‐loop receptors is conserved between prokaryotes and eukaryotes.     

Efficient DNA packaging of bacteriophage PRD1 requires the unique vertex protein P6

The assembly of bacteriophage PRD1 proceeds via formation of empty procapsids containing an internal lipid membrane, into which the linear double-stranded DNA genome is subsequently packaged. The packaging ATPase P9 and other putative packaging proteins have been shown to be located at a unique vertex of the PRD1 capsid. Here, we describe the isolation and characterization of a suppressor-sensitive PRD1 mutant deficient in the unique vertex protein P6. Protein P6 was found to be an essential part of the PRD1 packaging machinery; its absence leads to greatly reduced packaging efficiency. Lack of P6 was not found to affect particle assembly, because in the P6-deficient mutant infection, wild-type (wt) amounts of particles were produced, although most were empty. P6 was determined not to be a specificity factor, as the few filled particles seen in the P6-deficient infection contained only PRD1-specific DNA. The presence of P6 was not necessary for retention of DNA in the capsid once packaging had occurred, and P6-deficient DNA-containing particles were found to be stable and infectious, albeit not as infectious as wt PRD1 virions. A packaging model for bacteriophage PRD1, based on previous results and those obtained in this study, is presented.     

Mechanical stress alters protein O‐GlcNAc in human periodontal ligament cells

Protein O‐linked N‐acetylglucosamine (O‐GlcNAc) is a post‐translational modification of intracellular proteins that regulates several physiological and pathophysiological process, including response to various stressors. However, O‐GlcNAc's response to mechanical stress has not been investigated yet. As human periodontal ligament (PDL) cells are stimulated by compression force during orthodontic tooth movement that results in structural remodelling, in this study we investigated whether mechanical stress induces any alteration in protein O‐GlcNAc in PDL cells. In this study, PDL cells isolated from premolars extracted for orthodontic indications were exposed to 0, 1.5, 3, 7 and 14 g/cm² compression forces for 12 hours. Cell viability was measured by flow cytometry, and protein O‐GlcNAc was analysed by Western blot. Cellular structure and intracellular distribution of O‐GlcNAc was studied by immunofluorescence microscopy. We found that between 1.5 and 3 g/cm² mechanical compression, O‐GlcNAc significantly elevated; however, at higher forces O‐GlcNAc level was not increased. We also found that intracellular localization of O‐GlcNAc proteins became more centralized under 2 g/cm² compression force. Our results suggest that structural changes stimulated by compression forces have a significant effect on the regulation of O‐GlcNAc; thus, it might play a role in the mechanical stress adaptation of PDL cells.     

Phylogeography of the Temminck’s Stint (Calidris temminckii): historical vicariance but little present genetic structure in a regionally endangered Palearctic wader

Aim We study the population differentiation and phylogeography of the Temminck’s Stint (Calidris temminckii). Specifically, we seek signs of past and present population size changes and dispersal events and evaluate management and conservation unit status of the populations. We also study the possibility of introgression as the origin of two mitochondrial DNA (mtDNA) lineages found and estimate the divergence time of the lineages. Location Northern Eurasia. Methods We analysed 583 bp of mtDNA control region domains I and II and 11 microsatellite loci from 13 localities throughout the breeding range. In addition, we used mitochondrial cytochrome c oxidase subunit I (COI), a barcoding gene, to search for signs of introgression. Results More population differentiation was found from microsatellites than from mtDNA, although differentiation was weak in both markers. Signs of past population growth were observed, in addition to more recent decline in some areas. Both control region and COI sequences revealed two maternal lineages coexisting in Fennoscandia and in north‐west Siberia. No signs of introgression were detected. Lineage divergence time was estimated to have occurred during the glacial periods of Pleistocene. Main conclusions Slight differences in mtDNA and microsatellite differentiation and diversity may reflect different features – such as the mutation rate and effective population size – of the markers used, or female‐biased dispersal pattern and high male site‐fidelity of the species. The coexistence of the two mitochondrial lineages is most likely a consequence of post‐glacial mixing of two refugial Pleistocene populations. Based on genetic information alone, global conservation concerns are not imminent. However, fast decline of a marginal Bothnian Bay population and the smallness and remoteness of a Central Yakutian population warrant conservation actions.