Friedreich's Ataxia (GAA)n•(TTC)n Repeats Strongly Stimulate Mitotic Crossovers in Saccharomyces cerevisae

Expansions of trinucleotide GAA•TTC tracts are associated with the human disease Friedreich's ataxia, and long GAA•TTC tracts elevate genome instability in yeast. We show that tracts of (GAA)230•(TTC)230 stimulate mitotic crossovers in yeast about 10,000-fold relative to a “normal” DNA sequence; (GAA)n•(TTC)n tracts, however, do not significantly elevate meiotic recombination. Most of the mitotic crossovers are associated with a region of non-reciprocal transfer of information (gene conversion). The major class of recombination events stimulated by (GAA)n•(TTC)n tracts is a tract-associated double-strand break (DSB) that occurs in unreplicated chromosomes, likely in G1 of the cell cycle. These findings indicate that (GAA)n•(TTC)n tracts can be a potent source of loss of heterozygosity in yeast

Helicobacter pylori Perturbs Iron Trafficking in the Epithelium to Grow on the Cell Surface

Helicobacter pylori (Hp) injects the CagA effector protein into host epithelial cells and induces growth factor-like signaling, perturbs cell-cell junctions, and alters host cell polarity. This enables Hp to grow as microcolonies adhered to the host cell surface even in conditions that do not support growth of free-swimming bacteria. We hypothesized that CagA alters host cell physiology to allow Hp to obtain specific nutrients from or across the epithelial barrier. Using a polarized epithelium model system, we find that isogenic ΔcagA mutants are defective in cell surface microcolony formation, but exogenous addition of iron to the apical medium partially rescues this defect, suggesting that one of CagA's effects on host cells is to facilitate iron acquisition from the host. Hp adhered to the apical epithelial surface increase basolateral uptake of transferrin and induce its transcytosis in a CagA-dependent manner. Both CagA and VacA contribute to the perturbation of transferrin recycling, since VacA is involved in apical mislocalization of the transferrin receptor to sites of bacterial attachment. To determine if the transferrin recycling pathway is involved in Hp colonization of the cell surface, we silenced transferrin receptor expression during infection. This resulted in a reduced ability of Hp to colonize the polarized epithelium. To test whether CagA is important in promoting iron acquisition in vivo, we compared colonization of Hp in iron-replete vs. iron-deficient Mongolian gerbils. While wild type Hp and ΔcagA mutants colonized iron-replete gerbils at similar levels, ΔcagA mutants are markedly impaired in colonizing iron-deficient gerbils. Our study indicates that CagA and VacA act in concert to usurp the polarized process of host cell iron uptake, allowing Hp to use the cell surface as a replicative niche

UVA/UVA1 phototherapy and PUVA photochemotherapy in connective tissue diseases and related disorders: a research based review

BACKGROUND: Broad-band UVA, long-wave UVA1 and PUVA treatment have been described as an alternative/adjunct therapeutic option in a number of inflammatory and malignant skin diseases. Nevertheless, controlled studies investigating the efficacy of UVA irradiation in connective tissue diseases and related disorders are rare. METHODS: Searching the PubMed database the current article systematically reviews established and innovative therapeutic approaches of broad-band UVA irradiation, UVA1 phototherapy and PUVA photochemotherapy in a variety of different connective tissue disorders. RESULTS: Potential pathways include immunomodulation of inflammation, induction of collagenases and initiation of apoptosis. Even though holding the risk of carcinogenesis, photoaging or UV-induced exacerbation, UVA phototherapy seems to exhibit a tolerable risk/benefit ratio at least in systemic sclerosis, localized scleroderma, extragenital lichen sclerosus et atrophicus, sclerodermoid graft-versus-host disease, lupus erythematosus and a number of sclerotic rarities. CONCLUSIONS: Based on the data retrieved from the literature, therapeutic UVA exposure seems to be effective in connective tissue diseases and related disorders. However, more controlled investigations are needed in order to establish a clear-cut catalogue of indications

As cold as a fish? Relationships between the Dark Triad personality traits and affective experience during the day: A day reconstruction study

The Dark Triad of personality is a cluster of three socially aversive personality traits: Machiavellianism, narcissism and psychopathy. These traits are associated with a selfish, aggressive and exploitative interpersonal strategy. The objective of the current study was to establish relationships between the Dark Triad traits (and their dimensions) and momentary affect. Machiavellianism, grandiose narcissism, vulnerable narcissism and the dimensions of the Triarchic model of psychopathy (namely, boldness, meanness and disinhibition) were examined. We used the Day Reconstruction Method, which is based on reconstructing affective states experienced during the previous day. The final sample consisted of 270 university students providing affective ratings of 3047 diary episodes. Analyses using multilevel modelling showed that only boldness had a positive association with positive affective states and affect balance, and a negative association with negative affective states. Grandiose narcissism and its sub-dimensions had no relationship with momentary affect. The other dark traits were related to negative momentary affect and/or inversely related to positive momentary affect and affect balance. As a whole, our results empirically demonstrated distinctiveness of the Dark Triad traits in their relationship to everyday affective states. These findings are not congruent with the notion that people with the Dark Triad traits, who have a dispositional tendency to manipulate and exploit others, are generally cold and invulnerable to negative feelings. The associations between the Dark Triad and momentary affect were discussed in the contexts of evolutionary and positive psychology, in relation to the role and adaptive value of positive and negative emotions experienced by individuals higher in Machiavellianism, narcissism and psychopathy

Methods for assessing DNA repair and repeat expansion in Huntington's Disease

Huntington’s disease (HD) is caused by a CAG repeat expansion in the HTT gene. Repeat length can change over time, both in individual cells and between generations, and longer repeats may drive pathology. Cellular DNA repair systems have long been implicated in CAG repeat instability but recent genetic evidence from humans linking DNA repair variants to HD onset and progression has reignited interest in this area. The DNA damage response plays an essential role in maintaining genome stability, but may also license repeat expansions in the context of HD. In this chapter we summarize the methods developed to assay CAG repeat expansion/contraction in vitro and in cells, and review the DNA repair genes tested in mouse models of HD. While none of these systems is currently ideal, new technologies, such as long-read DNA sequencing, should improve the sensitivity of assays to assess the effects of DNA repair pathways in HD. Improved assays will be essential precursors to high-throughput testing of small molecules that can alter specific steps in DNA repair pathways and perhaps ameliorate expansion or enhance contraction of the HTT CAG repeat

The Wide-field Spectroscopic Telescope (WST) Science White Paper

The Wide-field Spectroscopic Telescope (WST) is proposed as a new facility dedicated to the efficient delivery of spectroscopic surveys. This white paper summarises the initial concept as well as the corresponding science cases. WST will feature simultaneous operation of a large field-of-view (3 sq. degree), a high multiplex (20,000) multi-object spectrograph (MOS) and a giant 3x3 sq. arcmin integral field spectrograph (IFS). In scientific capability these requirements place WST far ahead of existing and planned facilities. Given the current investment in deep imaging surveys and noting the diagnostic power of spectroscopy, WST will fill a crucial gap in astronomical capability and work synergistically with future ground and space-based facilities. This white paper shows that WST can address outstanding scientific questions in the areas of cosmology; galaxy assembly, evolution, and enrichment, including our own Milky Way; origin of stars and planets; time domain and multi-messenger astrophysics. WST's uniquely rich dataset will deliver unforeseen discoveries in many of these areas. The WST Science Team (already including more than 500 scientists worldwide) is open to the all astronomical community. To register in the WST Science Team please visit https://www.wstelescope.com/for-scientists/participat

Experimenting with Trinucleotide Repeats: Facts and Technical Issues

Part of the Methods in Molecular Biology book series (MIMB, volume 2056)International audienceTrinucleotide repeats are a peculiar class of microsatellites involved in many neurological as well as developmental disorders. Their propensity to generate very large expansions over time is supposedly due to their capacity to form specific secondary structures, such as imperfect hairpins, triple helices, or G-quadruplexes. These unusual structures were proposed to trigger expansions in vivo. Here, I review known technical issues linked to these structures, such as slippage during polymerase chain reaction and aberrant migration of long trinucleotide repeats during agarose gel electrophoresis. Our current understanding of interactions between trinucleotide repeat secondary structures and the mismatch-repair machinery is also quickly reviewed, and critical questions relevant to these interactions are addressed

Recombination-induced CAG trinucleotide repeat expansions in yeast involve the MRE11–RAD50–XRS2 complex

Recombination induced by double-strand breaks (DSBs) in yeast leads to a higher proportion of expansions to contractions than does replication-associated tract length changes. Expansions are apparently dependent on the property of the repeat array to form hairpins, since DSB repair of a CAA(87) repeat induces only contractions of the repeat sequence. DSB-repair efficiency is reduced by 40% when DNA synthesis must traverse a CAG(98) array, as compared with a CAA(87) array. These data indicate that repair- associated DNA synthesis is inhibited by secondary structures formed by CAG(98) and that these structures promote repeat expansions during DSB repair. Overexpression of Mre11p or Rad50p suppresses the inhibition of DSB repair by CAG(98) and significantly increases the average size of expansions found at the recipient locus. Both effects are dependent on the integrity of the Mre11p–Rad50p–Xrs2p complex. The Mre11 complex thus appears to be directly involved in removing CAG or CTG hairpins that arise frequently during DNA synthesis accompanying gene conversion of these trinucleotide repeats

Manufacture of a human mesenchymal stem cell population using an automated cell culture platform

Tissue engineering and regenerative medicine are rapidly developing fields that use cells or cell-based constructs as therapeutic products for a wide range of clinical applications. Efforts to commercialise these therapies are driving a need for capable, scaleable, manufacturing technologies to ensure therapies are able to meet regulatory requirements and are economically viable at industrial scale production. We report the first automated expansion of a human bone marrow derived mesenchymal stem cell population (hMSCs) using a fully automated cell culture platform. Differences in cell population growth profile, attributed to key methodological differences, were observed between the automated protocol and a benchmark manual protocol. However, qualitatively similar cell output, assessed by cell morphology and the expression of typical hMSC markers, was obtained from both systems. Furthermore, the critical importance of minor process variation, e.g. the effect of cell seeding density on characteristics such as population growth kinetics and cell phenotype, was observed irrespective of protocol type. This work highlights the importance of careful process design in therapeutic cell manufacture and demonstrates the potential of automated culture for future optimisation and scale up studies required for the translation of regenerative medicine products from the laboratory to the clinic