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Hypoxia-inducible factor (HIF) is the major transcriptional regulator of cellular responses to hypoxia. The two principal HIF-α isoforms, HIF-1α and HIF-2α, are progressively stabilized in response to hypoxia and form heterodimers with HIF-1β to activate a broad range of transcriptional responses. Here, we report on the pan-genomic distribution of isoform-specific HIF binding in response to hypoxia of varying severity and duration, and in response to genetic ablation of each HIF-α isoform. Our findings reveal that, despite an identical consensus recognition sequence in DNA, each HIF heterodimer loads progressively at a distinct repertoire of cell-type-specific sites across the genome, with little evidence of redistribution under any of the conditions examined. Marked biases towards promoter-proximal binding of HIF-1 and promoter-distant binding of HIF-2 were observed under all conditions and were consistent in multiple cell type. The findings imply that each HIF isoform has an inherent property that determines its binding distribution across the genome, which might be exploited to therapeutically target the specific transcriptional output of each isoform independently., (© 2018 The Authors. Published under the terms of the CC BY 4.0 license.)

Interactions between biological pathways and molecular oxygen require robust mechanisms for detecting and responding to changes in cellular oxygen availability, to support oxygen homeostasis. Peptidylglycine α-amidating monooxygenase (PAM) catalyzes a two-step reaction resulting in the C-terminal amidation of peptides, a process important for their stability and biological activity. Here we show that in human, mouse, and insect cells, peptide amidation is exquisitely sensitive to hypoxia. Different amidation events on chromogranin A, and on peptides processed from proopiomelanocortin, manifest similar striking sensitivity to hypoxia in a range of neuroendocrine cells, being progressively inhibited from mild (7% O2) to severe (1% O2) hypoxia. In developing Drosophila melanogaster larvae, FMRF amidation in thoracic ventral (Tv) neurons is strikingly suppressed by hypoxia. Our findings have thus defined a novel monooxygenase-based oxygen sensing mechanism that has the capacity to signal changes in oxygen availability to peptidergic pathways., (© 2015 by The American Society for Biochemistry and Molecular Biology, Inc.)

Medical management of endometriosis and fibroids involves manipulation of the hypothalamic-pituitary-gonadal axis to alter the balance of sex hormones thereby inhibiting disease progression and ameliorate symptoms. Unfortunately, resultant menopausal symptoms sometimes limit the tolerability and duration of such treatment. The use of gonadotrophin-releasing hormone agonists to treat these diseases can result in short-term hypoestrogenic and vasomotor side effects as well as long-term impacts on bone health and cardiovascular risk. The routine use of add-back hormone replacement has reduced these risks and increased patient compliance, making this group of drugs more useful as a medium-term treatment option. The estrogen threshold hypothesis highlights the concept of a 'therapeutic window' in which bone loss is minimal but the primary disease is not aggravated. It explains why add-back therapy is appropriate for such patients and helps to explain the basis behind new developments in the treatment of hormonally responsive gynaecological conditions such as gonadotrophin-releasing hormone antagonists and progesterone receptor modulators., (© The Author(s) 2015 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav.)

A large body of functional and epidemiological evidence have previously illustrated the impact of specific MHC class I subtypes on clinical outcome during HIV-1 infection, and these observations have recently been re-iterated in genome wide association studies (GWAS). Yet because of the complexities surrounding GWAS-based approaches and the lack of knowledge relating to the identity of rarer single nucleotide polymorphism (SNP) variants, it has proved difficult to discover independent causal variants associated with favourable immune control. This is especially true of the candidate variants within the HLA region where many of the recently proposed disease influencing SNPs appear to reflect linkage with 'protective' MHC class I alleles. Yet causal MHC-linked SNPs may exist but remain overlooked owing to the complexities associated with their identification. Here we focus on the ancestral TNFα promoter -237A variant (rs361525), shown historically to be in complete linkage disequilibrium with the 'protective' HLA-B*5701 allele. Many of the ancestral SNPs within the extended TNFα promoter have been associated with both autoimmune conditions and disease outcomes, however, the direct role of these variants on TNFα expression remains controversial. Yet, because of the important role played by TNFα in HIV-1 infection, and given the proximity of the -237 SNP to the core promoter, its location within a putative repressor region previously characterized in mice, and its disruption of a methylation-susceptible CpG dinucleotide motif, we chose to carefully evaluate its impact on TNFα production. Using a variety of approaches we now demonstrate that carriage of the A SNP is associated with lower TNFα production, via a mechanism not readily explained by promoter methylation nor the binding of transcription factors or repressors. We propose that the -237A variant could represent a minor causal SNP that additionally contributes to the HLA-B*5701-mediated 'protective' effect during HIV-1 infection.

Nox5 belongs to the calcium-regulated subfamily of NADPH oxidases (Nox). Like other calcium-regulated Noxes, Nox5 has an EF-hand-containing calcium-binding domain at its N-terminus, a transmembrane heme-containing region, and a C-terminal dehydrogenase (DH) domain that binds FAD and NADPH. While Nox1-4 require regulatory subunits, including p22phox, Nox5 activity does not depend on any subunits. We found that inactive point mutants and truncated forms of Nox5 (including the naturally expressed splice form, Nox5S) inhibit full-length Nox5, consistent with formation of a dominant negative complex. Oligomerization of full-length Nox5 was demonstrated using co-immunoprecipitation of coexpressed, differentially tagged forms of Nox5 and occurred in a manner independent of calcium ion. Several approaches were used to show that the DH domain mediates oligomerization: Nox5 could be isolated as a multimer when the calcium-binding domain and/or the N-terminal polybasic region (PBR-N) was deleted, but deletion of the DH domain eliminated oligomerization. Further, a chimera containing the transmembrane domain of Ciona intestinalis voltage sensor-containing phosphatase (CiVSP) fused to the Nox5 DH domain formed a co-immunoprecipitating complex with, and functioned as a dominant inhibitor of, full-length Nox5. Radiation inactivation of Nox5 overexpressed in HEK293 cells and endogenously expressed in human aortic smooth muscle cells indicated molecular masses of ∼350 and ∼300 kDa, respectively, consistent with a tetramer being the functionally active unit. Thus, Nox5 forms a catalytically active oligomer in the membrane that is mediated by its dehydrogenase domain. As a result of oligomerization, the short, calcium-independent splice form, Nox5S, may function as an endogenous inhibitor of calcium-stimulated ROS generation by full-length Nox5.

Digital Image-based Elasto Tomography (DIET) is a non-invasive breast cancer screening modality that induces vibrations into a breast and images its surface motion with digital cameras. Disturbances in the motion are caused by areas of higher stiffness within the breast, potentially cancerous tumors. A concept is presented to detect the angular location of a tumor by analyzing the phase delay of the vibrations on the surface. The approach is verified experimentally on silicone phantom breasts with stiffer inclusions ranging from 0-32 mm. A strong signal differentiating healthy and cancerous phantoms can be seen at the second modal frequency of the breast, clearly detecting a 10 mm tumor. This approach offers great potential for this low cost and accessible breast cancer screening, as an adjunct to existing modalities.

This paper reports the use of diodes as a quality control tool on more than 95% of patients undergoing megavoltage radiotherapy during the past 2 years. The paper describes a simple method for using the diodes, a number of possible pitfalls and a quality issue raised by the measurements. In general, the results have been reassuring with very few patients falling outside a +/- 5% action level.

A survey of radiographic technique and estimated entrance surface dose has been carried out for 364 chest radiographs performed with mobile X-ray equipment in the Intensive Therapy Unit (ITU) and 30 wards at Aberdeen Royal Infirmary. Data for these two types of location were compared, as were those for two film/screen systems used on the wards. Image quality assessments were made on sets of radiographs for two patients. Entrance skin doses for chest radiographs performed in the ITU were 50% greater than on the wards with the same film/screen system. The main technique difference was the use of shorter focus-to-skin distances (FSDs) in ITU. Doses with the Kodak Insight system were 20% higher than those using Du Pont Quanta III in similar locations. No correlation was found between image quality and entrance surface dose (ESD). Results from the survey were used to recommend exposure factors for shorter FSDs. A follow-up study revealed a 35-45% reduction in ESD for Kodak Insight and a 20% reduction for Quanta III.