Regulation of Glutamine Carrier Proteins by RNF5 Determines Breast Cancer Response to ER Stress-Inducing Chemotherapies

SummaryMany tumor cells are fueled by altered metabolism and increased glutamine (Gln) dependence. We identify regulation of the L-glutamine carrier proteins SLC1A5 and SLC38A2 (SLC1A5/38A2) by the ubiquitin ligase RNF5. Paclitaxel-induced ER stress to breast cancer (BCa) cells promotes RNF5 association, ubiquitination, and degradation of SLC1A5/38A2. This decreases Gln uptake, levels of TCA cycle components, mTOR signaling, and proliferation while increasing autophagy and cell death. Rnf5-deficient MMTV-PyMT mammary tumors were less differentiated and showed elevated SLC1A5 expression. Whereas RNF5 depletion in MDA-MB-231 cells promoted tumorigenesis and abolished paclitaxel responsiveness, SLC1A5/38A2 knockdown elicited opposing effects. Inverse RNF5hi/SLC1A5/38A2lo expression was associated with positive prognosis in BCa. Thus, RNF5 control of Gln uptake underlies BCa response to chemotherapies

Plant lectins: the ties that bind in root symbiosis and plant defense

Lectins are a diverse group of carbohydrate-binding proteins that are found within and associated with organisms from all kingdoms of life. Several different classes of plant lectins serve a diverse array of functions. The most prominent of these include participation in plant defense against predators and pathogens and involvement in symbiotic interactions between host plants and symbiotic microbes, including mycorrhizal fungi and nitrogen-fixing rhizobia. Extensive biological, biochemical, and molecular studies have shed light on the functions of plant lectins, and a plethora of uncharacterized lectin genes are being revealed at the genomic scale, suggesting unexplored and novel diversity in plant lectin structure and function. Integration of the results from these different types of research is beginning to yield a more detailed understanding of the function of lectins in symbiosis, defense, and plant biology in general

Expression of MsLEC1 transgenes in alfalfa plants causes symbiotic abnormalities

Legume lectins have been proposed to have important symbiotic roles during Rhizobium-legume symbioses. To test this hypothesis, the symbiotic responses of transgenic alfalfa plants that express a portion of the putative alfalfa lectin gene MsLEC1 or MsLEC2 in either the antisense or sense orientation were analyzed following inoculation with wild-type Sinorhizobium meliloti 1021. MsLEC1-antisense (LEC1AS) plants were stunted, exhibited hypernodulation, and developed not only abnormally large nodules but also numerous small nodules, both of which senesced prematurely. MsLEC2-antisense plants were intermediate in growth and nodule number compared with LEC1AS and vector control plants. The symbiotic abnormalities of MsLEC1-sense transgene plants were similar to but milder than the responses shown by the LEC1AS plants, whereas MsLEC2-sense transgene plants exhibited symbiotic responses that were identical to those of vector and nontransgenic control plants. MsLEC1 mRNA accumulation was not detected in nodule RNA by Northern blot analysis but was localized to alfalfa nodule meristems and the adjacent cells of the invasion zone by in situ hybridization; transcripts were also detected in root meristems. A similar spatial pattern of MsLEC2 expression was found by using a whole-mount in situ hybridization procedure. Moreover, mRNAs for an orthologous lectin gene (MaLEC) were detected in white sweetclover (Melilotus alba) nodules and root tips

Dimerization of Recombinant Tobacco Mosaic Virus Movement Protein

The p30 movement protein (MP) is essential for cell-to-cell spread of tobacco mosaic virus in planta. We used anion-exchange chromatography and preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) to obtain highly purified 30-kDa MP, which migrated as a single band in native PAGE. Analytical ultracentrifugation suggested that the protein was monodisperse and dimeric in the nonionic detergent n-octyl-β-d-glucopyranoside. Circular dichroism (CD) spectroscopy showed that the detergent-solubilized protein contained significant α-helical secondary structure. Proteolysis of the C-tail generated a trypsin-resistant core that was a mixture of primarily monomers and some dimers. We propose that MP dimers are stabilized by electrostatic interactions in the C terminus as well as hydrophobic interactions between putative transmembrane α-helical coiled coils

Systems analysis of the prostate tumor suppressor NKX3.1 supports roles in DNA repair and luminal cell differentiation [v2; ref status: indexed, http://f1000r.es/4x1]

NKX3.1 is a homeobox transcription factor whose function as a prostate tumor suppressor remains insufficiently understood because neither the transcriptional program governed by NKX3.1, nor its interacting proteins have been fully revealed. Using affinity purification and mass spectrometry, we have established an extensive NKX3.1 interactome which contains the DNA repair proteins Ku70, Ku80, and PARP, thus providing a molecular underpinning to previous reports implicating NKX3.1 in DNA repair. Transcriptomic profiling of NKX3.1-negative prostate epithelial cells acutely expressing NKX3.1 revealed a rapid and complex response that is a near mirror image of the gene expression signature of human prostatic intraepithelial neoplasia (PIN). Pathway and network analyses suggested that NKX3.1 actuates a cellular reprogramming toward luminal cell differentiation characterized by suppression of pro-oncogenic c-MYC and interferon-STAT signaling and activation of tumor suppressor pathways. Consistently, ectopic expression of NKX3.1 conferred a growth arrest depending on TNFα and JNK signaling. We propose that the tumor suppressor function of NKX3.1 entails a transcriptional program that maintains the differentiation state of secretory luminal cells and that disruption of NKX3.1 contributes to prostate tumorigenesis by permitting luminal cell de-differentiation potentially augmented by defects in DNA repair

NKX3.1 expression and interactions Dataset

<p>Data set 1: Summary of NKX3.1 interacting proteins</p> <p>Data set 1A: All proteins identified in four independent mock and FLAG-NKX3.1 affinity purifications<br>Data set 1B: List of 58 high confidence NKX3.1 interacting proteins<br>Data set 1C: Lists of proteins shown in the Venn diagram in Fig. 1B</p> <p>Data set 2: Summary of NKX3.1-regulated gene expression</p> <p>Data set 2A: All raw mRNA expression data<br>Data set 2B: Averaged mRNA expression data<br>Data set 2C: List of mRNAs that show a greater than 5-fold change (p greater or even to 0.05) in cells expressing NKX3.1 for 7 hours (= "5x data set")<br>Data set 2D: list of NKX3.1 regulated genes containing conserved AP1 binding sites<br>Data set 2E: List of NKX3.1 regulated genes containing conserved SRF binding sites<br>Data set 2F: List of NKX3.1 regulated mRNAs that are inversely regulated in human prostate cancer derived cell lines</p> <p> </p