Aquaporin-7 Regulates the Response to Cellular Stress in Breast Cancer.

The complex yet interrelated connections between cancer metabolism, gene expression, and oncogenic driver genes have the potential to identify novel biomarkers and drug targets with prognostic and therapeutic value. Here we effectively integrated metabolomics and gene expression data from breast cancer mouse models through a novel unbiased correlation-based network analysis. This approach identified 35 metabolite and 34 gene hubs with the most network correlations. These hubs have prognostic value and are likely integral to tumor metabolism and breast cancer. The gene hub Aquaporin-7 (Aqp7), a water and glycerol channel, was identified as a novel regulator of breast cancer. AQP7 was prognostic of overall survival in patients with breast cancer. In mouse breast cancer models, reduced expression of Aqp7 caused reduced primary tumor burden and lung metastasis. Metabolomics and complex lipid profiling of cells and tumors with reduced Aqp7 revealed significantly altered lipid metabolism, glutathione metabolism, and urea/arginine metabolism compared with controls. These data identify AQP7 as a critical regulator of metabolic and signaling responses to environmental cellular stresses in breast cancer, highlighting AQP7 as a potential cancer-specific therapeutic vulnerability. SIGNIFICANCE: Aquaporin-7 is identified as a critical regulator of nutrient availability and signaling that responds to cellular stresses, making it an attractive therapeutic target in breast cancer. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/19/4071/F1.large.jpg.

The CXCL5/CXCR2 axis is sufficient to promote breast cancer colonization during bone metastasis.

Bone is one of the most common sites for metastasis across cancers. Cancer cells that travel through the vasculature and invade new tissues can remain in a non-proliferative dormant state for years before colonizing the metastatic site. Switching from dormancy to colonization is the rate-limiting step of bone metastasis. Here we develop an ex vivo co-culture method to grow cancer cells in mouse bones to assess cancer cell proliferation using healthy or cancer-primed bones. Profiling soluble factors from conditioned media identifies the chemokine CXCL5 as a candidate to induce metastatic colonization. Additional studies using CXCL5 recombinant protein suggest that CXCL5 is sufficient to promote breast cancer cell proliferation and colonization in bone, while inhibition of its receptor CXCR2 with an antagonist blocks proliferation of metastatic cancer cells. This study suggests that CXCL5 and CXCR2 inhibitors may have efficacy in treating metastatic bone tumors dependent on the CXCL5/CXCR2 axis.

Quantitative proteomics of heavy metal exposure in Arabidopsis thaliana reveals alterations in one-carbon metabolism enzymes upon exposure to zinc.

Plant zinc (Zn) homeostasis must be tightly regulated as the requirement for this micronutrient necessitates its uptake. However, excessive Zn can lead to toxicity and the plant must respond rapidly and effectively within its capacity to minimize damage. To detect mechanisms that may be important for coping with excess Zn we carried out a quantitative proteomics approach using 2D-DIGE to identify Zn-responsive proteins in microsomal fractions from leaves of 4day, 200µM Zn-treated, Arabidopsis thaliana plants. Of the eight proteins which showed significant changes in abundance in the Zn-treated samples and which met all of the selection criteria following statistical analysis, six were successfully identified by LC-MS/MS with 2 or more unique peptides. Three of the proteins were found to be involved in the one-carbon metabolism pathway; including glycine decarboxylase P protein, serine hydroxymethyltransferase (SHMT) and methionine synthase, all of which showed reduced abundance in the Zn-treated samples. Western blot analysis confirmed the decrease in SHMT, while changes in metal tolerance protein indicated plants were most likely actively sequestering Zn. Interestingly, excess Zn led to increased petiole length, a phenotype which may reflect the reduced levels of methionine, a key product of the one-carbon metabolism pathway. BIOLOGICAL SIGNIFICANCE: Metal contamination is becoming an increasingly common environmental problem. High levels of zinc can be found in certain soils naturally or as a result of long-term anthropogenic activity which leads to its accumulation; i.e. use of fertilizers or industrial waste. The study of metal tolerant plants, particularly those classified as hyperaccumulators has been driven by the potential use of these plants for bioremediation purposes. However, the effects of heavy metal exposure on sensitive plants and the different cellular processes that are affected have received significantly less attention. We are interested in identifying proteins in A. thaliana that are induced as a result of exposure to subtoxic levels of heavy metals with the aim of discovering novel participants in heavy metal stress and adaptation.

Zinc tolerance and accumulation in stable cell suspension cultures and in vitro regenerated plants of the emerging model plant Arabidopsis halleri (Brassicaceae).

Arabidopsis halleri is increasingly employed as a model plant for studying heavy metal hyperaccumulation. With the aim of providing valuable tools for studies on cellular physiology and molecular biology of metal tolerance and transport, this study reports the development of successful and highly efficient methods for the in vitro regeneration of A. halleri plants and production of stable cell suspension lines. Plants were regenerated from leaf explants of A. halleri via a three-step procedure: callus induction, somatic embryogenesis and shoot development. Efficiency of callus proliferation and regeneration depended on the initial callus induction media and was optimal in the presence of 1 mg L(-1) 2,4-dichlorophenoxyacetic acid, and 0.05 mg L(-1) benzylaminopurine. Subsequent shoot and root regeneration from callus initiated under these conditions reached levels of 100% efficiency. High friability of the callus supported the development of cell suspension cultures with minimal cellular aggregates. Characterization of regenerated plants and cell cultures determined that they maintained not only the zinc tolerance and requirement of the whole plant but also the ability to accumulate zinc; with plants accumulating up to 50.0 micromoles zinc g(-1) FW, and cell suspension cultures 30.9 micromoles zinc g(-1) DW. Together this work will provide the experimental basis for furthering our knowledge of A. halleri as a model heavy metal hyperaccumulating plant.

Identification of a crucial histidine involved in metal transport activity in the Arabidopsis cation/H+ exchanger CAX1.

In plants, yeast, and bacteria, cation/H+ exchangers (CAXs) have been shown to translocate Ca2+ and other metal ions utilizing the H+ gradient. The best characterized of these related transporters is the plant vacuolar localized CAX1. We have used site-directed mutagenesis to assess the impact of altering the seven histidine residues to alanine within Arabidopsis CAX1. The mutants were expressed in a Saccharomyces cerevisiae strain that is sensitive to Ca2+ and other metals. By utilizing a yeast growth assay, the H338A mutant was the only mutation that appeared to alter Ca2+ transport activity. The CAX1 His338 residue is conserved among various CAX transporters and may be located within a filter for cation selection. We proceeded to mutate His338 to every other amino acid residue and utilized yeast growth assays to estimate the transport properties of the 19 CAX mutants. Expression of 16 of these His338 mutants could not rescue any of the metal sensitivities. However, expression of H338N, H338Q, and H338K allowed for some growth on media containing Ca2+. Most interestingly, H338N exhibited increased tolerance to Cd2+ and Zn2+. Endomembrane fractions from yeast cells were used to measure directly the transport of H338N. Although the H338N mutant demonstrated 25% of the wild type Ca2+/H+ transport, it showed an increase in transport for both Cd2+ and Zn2+ reflected in a decrease in the Km for these substrates. This study provides insights into the CAX cation filter and novel mechanisms by which metals may be partitioned across membranes.