Cycling to work in London and inhaled dose of black carbon.

Modelling studies suggest that urban cycling is associated with an increased inhaled dose of fossil fuel-derived black carbon (BC). Using the amount of black material in airway macrophages as a marker of long-term inhaled BC, we sought to compare inhaled BC dose in London (UK) cyclists and non-cyclists. Airway macrophage carbon was assessed in 28 (58%) out of 48 healthy adults (14 cyclists and 14 non-cyclists) who attended for induced sputum. Short-term (24 h) exposure to BC was assessed on a representative working day in 27 out of 28 subjects. Serum interleukin (IL)-1ß, IL-2, IL-6, IL-8, granulocyte-macrophage colony-stimulating factor and tumour necrosis factor (TNF)-α were assessed in 26 out of the 28 subjects. Cyclists were found to have increased airway macrophage carbon when compared with non-cyclists (mean ± se 1.81 ± 0.21 versus 1.11 ± 0.07 µm(2); p<0.01). Short-term monitoring showed no difference in 24 h BC exposure between the two groups. However, cyclists were exposed to higher concentrations of BC during commuting (p<0.01). Airway macrophage carbon was associated with monitored commute BC (n=28; r=0.47, p<0.05). TNF-α was found to be increased in cyclists (p<0.05), but no other cytokines were increased. Commuting to work by bicycle in London is associated with increased long-term inhaled dose of BC. Whether cycling per se increases inhaled BC dose remains unclear.

A rude awakening. Interview by Caroline Swinburne.

A recent film highlighted the possibility of awareness under anaesthetic. While a rare occurrence, it is invariably distressing.

SCF(Pof1)-ubiquitin and its target Zip1 transcription factor mediate cadmium response in fission yeast.

Ubiquitin-dependent proteolysis regulates gene expression in many eukaryotic systems. Pof1 is an essential fission yeast F-box protein that is homologous to budding yeast Met30. Temperature-sensitive pof1 mutants display acute growth arrest with small cell size. Extragenic suppressor analysis identified Zip1, a bZIP (basic leucine zipper) transcription factor, as a target for Pof1. We show Zip1 is stabilized in pof1 mutants, Pof1 binds only phosphorylated forms of Zip1, and Zip1 is ubiquitylated in vivo, indicating that Zip1 is a substrate of SCF(Pof1). Genome-wide DNA microarray assay shows that many cadmium-induced genes are under the control of Zip1, suggesting Zip1 plays a role in cadmium response. Consistently, zip1 mutants are hypersensitive to cadmium and unlike wild type, lose cell viability under this stress. Intriguingly, cadmium exposure results in upregulation of Zip1 levels and leads wild-type cells to growth arrest with reduced cell size, reminiscent of pof1 phenotypes. Our results indicate that Zip1 mediates growth arrest in cadmium response, which is essential to maintain viability. Normally growing cells prevent this response through constitutive ubiquitylation and degradation of Zip1 via SCF(Pof1).

Molecular interactions of fission yeast Skp1 and its role in the DNA damage checkpoint.

Skp1 is a central component of the E3 ubiquitin ligase SCF (Skp1-Cullin-1-F-box). It forms an adapter bridge between Cullin-1 and the substrate-determining component, the F-box protein. In order to establish the role of Skp1, a temperature sensitive (ts) screen was carried out using mutagenic PCR (polymerase chain reaction) and 9 independent ts mutants were isolated. Mapping the mutated residues on the 3-D structure of human Skp1 suggested that the mutants would be compromised in binding to F-box proteins but not Cullin-1 (Pcu1). In order to assess the binding properties of ts Skp1, 12 F-box proteins and Pcu1 were epitope-tagged, and co-immunoprecipitation performed. This systematic analysis showed that ts Skp1 retains binding to Pcu1. However, binding to three specific F-box proteins, essential Pof1, Pof3 involved in maintaining genome integrity, and nonessential Pof10, was reduced. skp1ts cells exhibit a G2 cell cycle delay, which is attributable to activation of the DNA damage checkpoint. Intriguingly, contrary to pof3 mutants, in which this checkpoint is required for survival, checkpoint abrogation in skp1(ts) suppresses a G2 delay and furthermore almost rescues the ts phenotype. The activation mechanism of the DNA damage checkpoint therefore differs between pof3Delta and skp1(ts), implicating a novel role for Skp1 in the checkpoint-signalling cascade.

Requirement of the SCFPop1/Pop2 Ubiquitin Ligase for Degradation of the Fission Yeast S Phase Cyclin Cig2.

Two multiprotein E3 (ubiquitin-protein ligase) ubiquitin ligases, the SCF (Skp1-Cullin-1-F-box) and the APC/C (anaphase promoting complex/cyclosome), are vital in ensuring the temporal order of the cell cycle. Particularly, timely destruction of cyclins via these two E3s is essential for down-regulation of cyclin-dependent kinase. In general, G(1) and S phase cyclins are ubiquitylated by the SCF, whereas ubiquitylation of mitotic cyclins is catalyzed by the APC/C. Here we show that fission yeast S phase cyclin Cig2 is ubiquitylated and degraded via both the SCF and the APC/C. Cig2 instability during G(2) and M phase is dependent upon the SCF complex, whereas the APC/C is responsible for Cig2 destruction during anaphase and G(1), thereby ensuring a spike pattern of Cig2 levels, peaking only at S phase. Two F-box/WD proteins Pop1 and Pop2, homologues of budding yeast Cdc4 and human Fbw7, are responsible for Cig2 instability. Pop1 binds Cig2 in vivo. An in vitro binding assay shows that an internal 93 amino acid residues comprising a part of the cyclin box are necessary and sufficient for this binding. Cig2 phosphorylation is also required for interaction with Pop1. We previously showed that transcriptional oscillation of cig2(+) requires Pop1 and Pop2 function. SCF(Pop1/Pop2) therefore regulates Cig2 levels in a dual manner, transcriptionally and post-translationally. Our results also highlight a collaborative action of the APC/C and the SCF toward the common substrate Cig2. This type of composite degradation control may be more general as the regulatory mechanism in other complex systems.

The expectations, healt beliefs and behaviour of patients seeking homoeopathic and conventional medicine

Research was carried out to determine whether or not there was difference in the health beliefs, expectations and behaviour of a sample of 92 patients attending a dermatological or rheumatological outpatient clinic which offered either homoeopathic treatment (N=47) or conventional treatment (N=45). Self-administered questionnaires were used which examined patients'pathwaysto care, expectations, beliefs, behaviour and multidimensional health locus of control. The two key differences between those seeking homoeopathic and those seeking conventional medicine were in terms of their reasons for attending a homeopathic or conventional clinic, and their beliefs about their presenting dermatological or rheumatological condition