The absence of SIR2alpha protein has no effect on global gene silencing in mouse embryonic stem cells

The yeast sir2 gene plays a central role in mediating gene silencing and DNA repair in this organism. The mouse sir2alpha gene is closely related to its yeast homologue and encodes a nuclear protein expressed at particularly high levels in embryonic stem (ES) cells. We used homologous recombination to create ES cells null for sir2alpha and found that these cells did not have elevated levels of acetylated histones and did not ectopically express silent genes. Unlike yeast sir2 mutants, our sir2alpha null ES cells had normal sensitivity to insults such as ionizing radiation and heat shock, and they were able to silence invading retroviruses normally. These sir2alpha null cells were able to differentiate in culture normally. Our results failed to provide evidence that the mammalian SIR2alpha protein plays a role in gene silencing and suggest that the physiological substrate(s) for the SIR2alpha deacetylase may be nuclear proteins other than histones.     

Synthesis of mycothiol, 1D-1-O-(2-[N-acetyl-L-cysteinyl]amino-2-deoxy-alpha-D-glucopyranosyl)-myo-inositol, principal low molecular mass thiol in the actinomycetes

Members of the actinomycetes produce 1D-1-O-(2-[N-acetyl-L-cysteinyl]amino-2-deoxy-alpha-D-glucopyranosyl)-myo-inositol or mycothiol 1 as principal low molecular mass thiol. Chemical synthesis of a biosynthetic precursor of mycothiol, the pseudodisaccharide 1D-1-O-(2-amino-2-deoxy-alpha-D-glucopyranosyl)-myo-inositol 13 was achieved by the following steps: (1) Enantioselective synthesis gave the glycosyl acceptors (-)-2,3,4,5,6-penta-O-acetyl-D-myo-inositol D-7 and the corresponding L-isomer L-7. (2) Condensation of D-7 and L-7 with the glycosyl donor 3,4,6-tri-O-acetyl-2-deoxy-2-(2,4-dinitrophenylamino)-alpha-D-glucopyranosylbromide afforded the corresponding alpha and beta anomeric products, which could be resolved by silica gel chromatography. (3) Deprotection of these by hydrolysis using an anion exchange resin gave 1D- and 1L-1-O-(2-amino-2-deoxy-alpha-D-glucopyranosyl)-myo-inositol 13 and 15 and the corresponding beta-coupled anomers 14 and 16. Only 13, and to a much lesser extent 15, were used by enzymes present in an ammonium sulphate fraction of a cellfree extract of Mycobacterium smegmatis for the enzymatic synthesis of mycothiol. In the absence of acetyl-SCoA, the immediate biosynthetic precursor of 1, desacetylmycothiol, was the major product.     

Sources of error in neurotransmitter analysis

A compound having fluorescence characteristics similar to those of 5-hydroxytryptamine (5-HT), when reacted with ninhydrin, is present in several invertebrates (Fasciola hepatica, Aplysia californica, Tritonia diomedia, and Hirudo medicinalis). However, this substance is not identical with 5-HT [ Andreini, G. C., Beretta, C., Faustini, R., and Gallina, G. (1970)Experientia26, 166–167]. We confirmed these findings for Fasciola and also observed this substance in Spirometra mansonoides and mouse brain. Using chromatography and amino acid analysis we identified this substance as lysine. While 5-HT is also demonstrable in Spirometra, no 5-HT is present in Fasciola. Although epinephrine and dopamine seem to be present in Spirometra when the hydroxyindole technique of Laverty and Taylor [ Laverty, R., and Taylor, K. M. (1968)Anal. Biochem.22, 269–279] is used, neither amine is detectable in adult Spirometra by mass spectrometric analysis. A relatively high concentration of tyrosine in Spirometra could account for the apparent presence of dopamine. Therefore, lysine, tyrosine, and possibly other substances can be sources of error in the detection of biogenic amines in invertebrates.     

Role of φ29 connector channel loops in late-stage DNA packaging

Double-stranded DNA bacteriophages and their eukaryotic virus counterparts have 12-fold head-tail connector assemblages embedded at a unique capsid vertex. This vertex is the site of assembly of the DNA packaging motor, and the connector has a central channel through which viral DNA passes during genome packaging and subsequent host infection. Crystal structures of connectors from different phages reveal either disordered residues or structured loops that project into the connector channel. Given the proximity to the translocating DNA substrate, these loops have been proposed to play a role in DNA packaging. Previous models have proposed structural motions in either the packaging ATPase or the connector channel loops as the driving force that translocates the DNA into the prohead. Here, we mutate the channel loops of the Bacillus subtilis bacteriophage φ29 connector and show that these loops have no active role in translocation of DNA. Instead, they appear to have an essential function near the end of packaging, acting to retain the packaged DNA in the head in preparation for motor detachment and subsequent tail assembly and virion completion.     

Environmental Factors Associated with the Carriage of Bacterial Pathogens in Norway Rats

Worldwide, Norway rats (Rattus norvegicus) carry a number of zoonotic pathogens. Many studies have identified rat-level risk factors for pathogen carriage. The objective of this study was to examine associations between abundance, microenvironmental and weather features and Clostridium difficile, antimicrobial-resistant (AMR) Escherichia coli and methicillin-resistant Staphylococcus aureus (MRSA) carriage in urban rats. We assessed city blocks for rat abundance and 48 microenvironmental variables during a trap-removal study, then constructed 32 time-lagged temperature and precipitation variables and fitted multivariable logistic regression models. The odds of C. difficile positivity were significantly lower when mean maximum temperatures were high (≥ 12.89°C) approximately 3 months before rat capture. Alley pavement condition was significantly associated with AMR E. coli. Rats captured when precipitation was low (< 49.40 mm) in the 15 days before capture and those from blocks that contained food gardens and institutions had increased odds of testing positive for MRSA. Different factors were associated with each pathogen, which may reflect varying pathogen ecology including exposure and environmental survival. This study adds to the understanding of how the microenvironment and weather impacts the epidemiology and ecology of zoonotic pathogens in urban ecosystems, which may be useful for surveillance and control activities.     

Erythrocyte spectrin is an E2 ubiquitin conjugating enzyme

The involvement of red blood cell spectrin in the ubiquitination process was studied. Spectrin was found to form two ubiquitin-associated derivatives, a DTT-sensitive ubiquitin adduct and a DTT-insensitive conjugate, characteristic intermediate and final products of the ubiquitination reaction cascade. In addition to spectrin and ubiquitin, ubiquitin-activating enzyme (E1) and ATP were necessary and sufficient to form both the spectrin-ubiquitin adduct and conjugate. No exogenous ubiquitin-conjugating (E2) or ligase (E3) activities were required, suggesting that erythrocyte spectrin is an E2 ubiquitin-conjugating enzyme able to target itself. Both ubiquitin adduct and conjugate were linked to the alpha subunit of spectrin, suggesting that the ubiquitin-conjugating (UBC) domain and its target regions reside on the same subunit.     

Ethnobotanical plant uses in the KwaNibela Peninsula, St Lucia, South Africa

Ethnobotanical field studies were conducted for the first time in the KwaNibela Peninsula of southern Maputaland, KwaZulu-Natal, to document indigenous knowledge about useful plants. The vernacular names and uses of 82 plant species were recorded and compared to published Zulu and Swazi knowledge. Medicines for skin disorders, toothache, wounds, worms, chest and throat ailments, infertility and purgatives are still commonly used. Superstition and divination play a major role in the traditional knowledge system of the people of KwaNibela with 24 plants used for this purpose. Three KwaNibela medicinal plants (Erythroxylum delagoense, Putterlickia verrucosa, and Teclea natalensis) appear to be new records, not previously reported in the general scientific literature. The list also includes 61 novel uses of plants and another 15 new variations on known (published) uses. Ten previously unpublished vernacular names are presented, together with an additional 19 new variants of known names. These new additions to the scientific literature confirm that indigenous knowledge in KwaZulu-Natal is not yet completely recorded.