The ZntA-like NpunR4017 plays a key role in maintaining homeostatic levels of zinc in Nostoc punctiforme

Analysis of cellular response to zinc exposure provides insights into how organisms maintain homeostatic levels of zinc that are essential, while avoiding potentially toxic cytosolic levels. Using the cyanobacterium Nostoc punctiforme as a model, qRT-PCR analyses established a profile of the changes in relative mRNA levels of the ZntA-like zinc efflux transporter NpunR4017 in response to extracellular zinc. In cells treated with 18 μM of zinc for 1 h, NpunR4017 mRNA levels increased by up to 1300 % above basal levels. The accumulation and retention of radiolabelled (65)Zn by NpunR4107-deficient and overexpressing strains were compared to wild-type levels. Disruption of NpunR4017 resulted in a significant increase in zinc accumulation up to 24 % greater than the wild type, while cells overexpressing NpunR4107 accumulated 22 % less than the wild type. Accumulation of (65)Zn in ZntA(-) Escherichia coli overexpressing NpunR4017 was reduced by up to 21 %, indicating the capacity for NpunR4017 to compensate for the loss of ZntA. These findings establish the newly identified NpunR4017 as a zinc efflux transporter and a key transporter for maintaining zinc homeostasis in N. punctiforme

Characterization of two cation diffusion facilitators NpunF0707 and NpunF1794 in Nostoc punctiforme

AIMS: To characterize genes involved in maintaining homeostatic levels of zinc in the cyanobacterium Nostoc punctiforme. METHODS AND RESULTS: Metal efflux transporters play a central role in maintaining homeostatic levels of trace elements such as zinc. Sequence analyses of the N. punctiforme genome identified two potential cation diffusion facilitator (CDF) metal efflux transporters, Npun_F0707 (Cdf31) and Npun_F1794 (Cdf33). Deletion of either Cdf31or Cdf33 resulted in increased zinc retention over 3 h. Interestingly, Cdf31(-) and Cdf33(-) mutants showed no change in sensitivity to zinc exposure in comparison with the wild type, suggesting some compensatory capacity for the loss of each other. Using qRT-PCR, a possible interaction was observed between the two cdf\u27s, where the Cdf31(-) mutant had a more profound effect on cdf33 expression than Cdf33(-) did on cdf31. Over-expression of Cdf31 and Cdf33 in ZntA(-) - and ZitB(-) -deficient Escherichia coli revealed function similarities between the ZntA and ZitB of E. coli and the cyanobacterial transporters. CONCLUSIONS: The data presented shed light on the function of two important transporters that regulate zinc homeostasis in N. punctiforme. SIGNIFICANCE AND IMPACT OF THE STUDY: This study shows for the first time the functional characterization of two cyanobacterial zinc efflux proteins belonging to the CDF family

PMC42, a novel model for the differentiated human breast

Cultured human breast carcinoma cell lines are important models for investigating the pathogenesis of breast cancer. Their use, however, is limited because of loss of expression of breast-specific markers and the development of a dedifferentiated phenotype after continuous culture. PMC42 is a unique human breast carcinoma line, previously shown to express secretory and myoepithelial markers. We have induced PMC42 cells to form hollow organoids in culture, similar to in vivo breast structures, using a combination of hormones including estrogen, progesterone, dexamethasone, insulin, and prolactin in combination with a permeable extracellular matrix. The organoids comprised polarized cells located around a central lumen. Expression of &beta;-casein was demonstrated in cells within organoids using reverse transcriptase-polymerase chain reaction, Western blot analysis, and confocal immunofluorescence. In this in vitro system, milk-specific gene expression was induced through hormone and matrix interactions which may be similar to those operating in vivo. PMC42 is a novel model for investigations into the molecular mechanisms of carcinogenesis and differentiation in the human breast. <br /

A new strategy for vascular complications in young people with type 1 diabetes mellitus

Diabetes vascular complications, including cardiovascular disease, diabetic nephropathy and retinopathy, have a negative effect on the long-term prognosis of young people with type 1 diabetes mellitus (T1DM). Poor glycaemic control and consequent increased HbA1c levels are major risk factors for the development of vascular complications. HbA1c levels are the main focus of current management strategies; however, the recommended target is rarely achieved in adolescents. Thus, a clear need exists for improved biomarkers to identify high-risk young people early and to develop new intervention strategies. Evidence is accumulating that early increases in urinary albumin excretion could be predictive of adolescents with T1DM who are at an increased risk of developing vascular complications, independent of HbA1c levels. These findings present an opportunity to move towards the personalized care of adolescents with T1DM, which takes into consideration changes in albumin excretion and other risk factors in addition to HbA1c levels.M. Loredana Marcovecchio, R. Neil Dalton, Denis Daneman, John Deanfield, Timothy W. Jones, H. Andrew W. Neil ... et al. (on behalf of the Adolescent type 1 Diabetes cardio-renal Intervention Trial (AdDIT) study group