Insertional mutagenesis to isolate acetate-requiring mutants in Chlamydomonas reinhardtii

Abstract An arg 7 mutant of the green alga Chlamydomonas reinhardtii was transformed with pARG7.8, a plasmid bearing the wild-type ARG 7 gene. Out of 4100 arg⁺ transformants selected on an arginine-free medium supplemented with acetate, nine failed to grow on acetate-free medium (ac⁻ mutants). The results of the genetic and molecular analysis of several ac⁻ mutants are in agreement with the hypothesis that they originated from insertion of the incoming plasmid into the nuclear genome. These mutants should constitute valuable tools for isolating the corresponding wild-type genes after plasmid rescue into Escherichia coli .     

Ring (15) chromosome

Summary Cytogenetic studies on lymphocytes from a girl aged 3 years and 10 months revealed a ring chromosome 15. Several banding methods showed the r(15) chromosome not to have any apparent deletion of the long arm. The silver staining technique for nucleolar organizer regions showed an NOR positive region (band p12). In only a few cells was a chromosome 15 missing. The size of the r(15) was found to be constant. Comparison with 11 previous reported cases in the literature shows that the clinical manifestations in the different patients with ring chromosome 15 are constant although not clinically identifiable and it appears likely to attribute them to a significantly retarded intrauterine and postnatal growth instead of presumed deficiency in the long arm and mosaic configurations.     

Measurements at theπE3 single beam correlated to dosimetry and therapy-planning for the pion applicator at SIN

Summary Using ionization chambers, aluminium activation, TLD and scintillation counters 3-dimensional total dose-distributions, stardose-distributions and pion stop-distributions have been measured in a single pion beam for various momenta and momentum spreads.It is demonstrated how these data will be used as an input into the therapy-planning program. The techniques developed are suited to check dynamical treatment with 60 pion beams.     

Functional interactions between coexisting toxin-antitoxin systems of the ccd family in Escherichia coli O157:H7

Toxin-antitoxin (TA) systems are widely represented on mobile genetic elements as well as in bacterial chromosomes. TA systems encode a toxin and an antitoxin neutralizing it. We have characterized a homolog of the ccd TA system of the F plasmid (ccd(F)) located in the chromosomal backbone of the pathogenic O157:H7 Escherichia coli strain (ccd(O157)). The ccd(F) and the ccd(O157) systems coexist in O157:H7 isolates, as these pathogenic strains contain an F-related virulence plasmid carrying the ccd(F) system. We have shown that the chromosomal ccd(O157) system encodes functional toxin and antitoxin proteins that share properties with their plasmidic homologs: the CcdB(O157) toxin targets the DNA gyrase, and the CcdA(O157) antitoxin is degraded by the Lon protease. The ccd(O157) chromosomal system is expressed in its natural context, although promoter activity analyses revealed that its expression is weaker than that of ccd(F). ccd(O157) is unable to mediate postsegregational killing when cloned in an unstable plasmid, supporting the idea that chromosomal TA systems play a role(s) other than stabilization in bacterial physiology. Our cross-interaction experiments revealed that the chromosomal toxin is neutralized by the plasmidic antitoxin while the plasmidic toxin is not neutralized by the chromosomal antitoxin, whether expressed ectopically or from its natural context. Moreover, the ccd(F) system is able to mediate postsegregational killing in an E. coli strain harboring the ccd(O157) system in its chromosome. This shows that the plasmidic ccd(F) system is functional in the presence of its chromosomal counterpart.     

Use of Electronic Loggers to Measure Changes in the Rates of Hand Washing with Soap in Low-Income Urban Households in India

We evaluated the utility of electronic loggers to measure the effects of a simple intervention designed to influence the rates of hand washing with soap within enclosed toilets and bathrooms in low-income urban households in Kerala, India. 58 households were given three items with embedded electronic loggers for a period of 2-5 days. Two logged soaps tracked hand and body washing in the bathroom. The third logged item was a water vessel used for flushing the toilet and for post-defecation anal cleansing; this served as a marker of toilet use. In addition, 28 households in a Soap by toilet arm were given an additional logged soap, to be kept by the toilet, and used for hand washing. Compared with the Soap in bathroom arm, the loggers in the Soap by toilet households recorded 73% greater daily use of soaps designated for hand washing (t(36)=2.92, p<0.01) and 172% greater use within 2 minutes of the use of the water vessel (t(36)=3.51, p = 0.001). We conclude that the loggers were capable of detecting changes in the rates of hand washing with soap and changes in hand washing with soap after use of the toilet. Further adoption of logger technologies would enable more insightful studies of hand washing within urban environments.     

Pre-clinical and clinical significance of heparanase in Ewing's sarcoma

Heparanase is an endoglycosidase that specifically cleaves heparan sulphate side chains of heparan sulphate proteoglycans, activity that is strongly implicated in cell migration and invasion associated with tumour metastasis, angiogenesis and inflammation. Heparanase up-regulation was documented in an increasing number of human carcinomas, correlating with reduced post-operative survival rate and enhanced tumour angiogenesis. Expression and significance of heparanase in human sarcomas has not been so far reported. Here, we applied the Ewing's sarcoma cell line TC71 and demonstrated a potent inhibition of cell invasion in vitro and tumour xenograft growth in vivo upon treatment with a specific inhibitor of heparanase enzymatic activity (compound SST0001, non-anticoagulant N-acetylated, glycol split heparin). Next, we examined heparanase expression and cellular localization by immunostaining of a cohort of 69 patients diagnosed with Ewing's sarcoma. Heparanase staining was noted in all patients. Notably, heparanase staining intensity correlated with increased tumour size (P = 0.04) and with patients' age (P = 0.03), two prognostic factors associated with a worse outcome. Our study indicates that heparanase expression is induced in Ewing's sarcoma and associates with poor prognosis. Moreover, it encourages the inclusion of heparanase inhibitors (i.e. SST0001) in newly developed therapeutic modalities directed against Ewing's sarcoma and likely other malignancies.     

Stress granules counteract senescence by sequestration of PAI‐1

Cellular senescence is a physiological response by which an organism halts the proliferation of potentially harmful and damaged cells. However, the accumulation of senescent cells over time can become deleterious leading to diseases and physiological decline. Our data reveal a novel interplay between senescence and the stress response that affects both the progression of senescence and the behavior of senescent cells. We show that constitutive exposure to stress induces the formation of stress granules (SGs) in proliferative and presenescent cells, but not in fully senescent cells. Stress granule assembly alone is sufficient to decrease the number of senescent cells without affecting the expression of bona fide senescence markers. SG‐mediated inhibition of senescence is associated with the recruitment of the plasminogen activator inhibitor‐1 (PAI‐1), a known promoter of senescence, to these entities. PAI‐1 localization to SGs increases the translocation of cyclin D1 to the nucleus, promotes RB phosphorylation, and maintains a proliferative, non‐senescent state. Together, our data indicate that SGs may be targets of intervention to modulate senescence in order to impair or prevent its deleterious effects.     

Deciphering CAPTCHAs: what a Turing test reveals about human cognition

Turning Turing's logic on its head, we used widespread letter-based Turing Tests found on the internet (CAPTCHAs) to shed light on human cognition. We examined the basis of the human ability to solve CAPTCHAs, where machines fail. We asked whether this is due to our use of slow-acting inferential processes that would not be available to machines, or whether fast-acting automatic orthographic processing in humans has superior robustness to shape variations. A masked priming lexical decision experiment revealed efficient processing of CAPTCHA words in conditions that rule out the use of slow inferential processing. This shows that the human superiority in solving CAPTCHAs builds on a high degree of invariance to location and continuous transforms, which is achieved during the very early stages of visual word recognition in skilled readers.