CLASSICAL PHOTOABSORPTION SUM RULES

We use a quantum loop expansion to derive sum rule constraints on polarized photoabsorption cross sections in the Standard Model, generalizing earlier results obtained by Altarelli, Cabibbo, and Maiani. We show that the logarithmic integral of the spin-dependent photoabsorption cross section $\int^\infty_{\nu_{th}} {d\nu\over \nu} \Delta \sigma_{\rm Born}(\nu)$ vanishes for any $2 \to 2$ Standard Model or supersymmetric process $\gamma a \to b c$ in the classical, tree-graph approximation. Here $\nu = {p \cdot q}/M$ and $\Delta \sigma(\nu) = \sigma_P(\nu)- \sigma_A(\nu)$ is the difference between the photoabsorption cross section for parallel and antiparallel photon and target helicities. Tests of the sum rule for the reactions $\gamma e \to W \nu$ and $\gamma \gamma \to W^+ W^-$ can provide new tests of the canonical magnetic and quadrupole couplings of the Standard Model. We also extend the sum rule to certain virtual photon processes.Comment: 11 pages, 1 figure. For hard copy, e-mail request for SLAC-PUB-95-6761 to [email protected]

A RasGAP SH3 Peptide Aptamer Inhibits RasGAP-Aurora Interaction and Induces Caspase-Independent Tumor Cell Death

The Ras GTPase-activating protein RasGAP catalyzes the conversion of active GTP-bound Ras into inactive GDP-bound Ras. However, RasGAP also acts as a positive effector of Ras and exerts an anti-apoptotic activity that is independent of its GAP function and that involves its SH3 (Src homology) domain. We used a combinatorial peptide aptamer approach to select a collection of RasGAP SH3 specific ligands. We mapped the peptide aptamer binding sites by performing yeast two-hybrid mating assays against a panel of RasGAP SH3 mutants. We examined the biological activity of a peptide aptamer targeting a pocket delineated by residues D295/7, L313 and W317. This aptamer shows a caspase-independent cytotoxic activity on tumor cell lines. It disrupts the interaction between RasGAP and Aurora B kinase. This work identifies the above-mentioned pocket as an interesting therapeutic target to pursue and points its cognate peptide aptamer as a promising guide to discover RasGAP small-molecule drug candidates

W ANOMALOUS MOMENTS AND THE POLARIZATION ASYMMETRY ZERO IN gamma e --> W nu

We show from general principles that there must be a center of mass energy, $\sqrt s_0$, where the polarization asymmetry $A=\Delta \sigma({\gamma e \to W \nu })/ \sigma({\gamma e \to W \nu })$ for circularly-polarized photon and electron beams vanishes. In the case of the Standard Model, the crossing point where the asymmetry changes sign occurs in Born approximation at $\sqrt s_{\gamma e} = 3.1583\ldots M_W \simeq 254$ GeV. We demonstrate the sensitivity of the position of the polarization asymmetry zero to modifications of the SM trilinear $\gamma W W$ coupling. Given reasonable assumptions for the luminosity and energy range for the Next Linear Collider(NLC) with a backscattered laser beam, we show that the zero point, $\sqrt s_0$, of the polarization asymmetry may be determined with sufficient precision to constrain the anomalous couplings of the $W$ to better than the 1\% level at $95\%$ CL. In addition to the fact that only a limited range of energy is required, the polarization asymmetry measurements have the important advantage that many of the systematic errors cancel in taking cross section ratios. The position of the zero thus provides an additional weapon in the arsenal used to probe anomalous trilinear gauge couplings.Comment: 21 pages, uuencoded postscript file. For hard copy, send e-mail to [email protected]

A universal transgene silencing method based on RNA interference

Inducible gene expression systems have contributed significantly to the understanding of molecular regulatory networks. Here we describe a simple and powerful RNA interference-based method that can silence the expression of any transgene. We first used an IRES bicistronic lentiviral vector and showed that targeting the second cistron with a specific siRNA resulted in silencing of both transgenes. We then inserted a siRNA minimal target sequence in the 3′-untranslated region (3′-UTR) of a transgene and showed that the cognate siRNA delivered by a lentiviral vector led to the partial silencing of the transgene. The multimerization of this siRNA target sequence led to the highly efficient silencing of four different transgenes. This new method to silence transgene expression is more versatile than existing methods of conditional inactivation of gene expression, such as transcriptional switches or site-specific recombination. It is applicable to a wide variety of models including primary cells, terminally differentiated cells and transgenic animals

Redox regulation of EGFR steers migration of hypoxic mammary cells towards oxygen

International audienceAerotaxis or chemotaxis to oxygen was described in bacteria 130 years ago. In eukaryotes, the main adaptation to hypoxia currently described relies on HIF transcription factors. To investigate whether aerotaxis is conserved in higher eukaryotes, an approach based on the self-generation of hypoxia after cell confinement was developed. We show that epithelial cells from various tissues migrate with an extreme directionality towards oxygen to escape hypoxia, independently of the HIF pathway. We provide evidence that, concomitant to the oxygen gradient, a gradient of reactive oxygen species (ROS) develops under confinement and that antioxidants dampen aerotaxis. Finally, we establish that in mammary cells, EGF receptor, the activity of which is potentiated by ROS and inhibited by hypoxia, represents the molecular target that guides hypoxic cells to oxygen. Our results reveals that aerotaxis is a property of higher eukaryotic cells and proceeds from the conversion of oxygen into ROS

PRMT1 Is Critical for the Transcriptional Activity and the Stability of the Progesterone Receptor.

International audienceThe progesterone receptor (PR) is an inducible transcription factor that plays critical roles in female reproductive processes and in several aspects of breast cancer tumorigenesis. Our report describes the type I protein arginine methyltransferase 1 (PRMT1) as a cofactor controlling progesterone pathway, through the direct methylation of PR. Mechanistic assays in breast cancer cells indicate that PRMT1 methylates PR at the arginine 637 and reduces the stability of the receptor, thereby accelerating its recycling and finally its transcriptional activity. Depletion of PRMT1 decreases the expression of a subset of progesterone-inducible genes, controlling breast cancer cells proliferation and migration. Consistently, Kaplan-Meier analysis revealed that low expression of PRMT1 predicts a longer survival among the subgroup with high PR. Our study highlights PR methylation as a molecular switch adapting the transcription requirement of breast cells during tumorigenesis

Combination of a discovery LC-MS/MS analysis and a label-free quantification for the characterization of an epithelial-mesenchymal transition signature

International audienceDisease phenotype reorganizations are the consequences of signaling pathway perturbations and protein abundance modulations. Characterizing the protein signature of a biological event allows the identification of new candidate biomarkers, new targets for treatments and selective patient therapy. The combination of discovery LC-MS/MS analyses and targeted mass spectrometry using selected reaction monitoring (SRM) mode has emerged as a powerful technology for biomarker identification and quantification owing to faster development time and multiplexing capability. The epithelial mesenchymal transition (EMT) is a process that controls local invasion and metastasis generation by stimulating changes in adhesion and migration of cells but also in metabolic pathways. In this study, the non-transformed human breast epithelial cell line MCF10A, treated by TGF beta or overexpressing mutant K-Ras(v12), two EMT inducers frequently involved in cancer progression, was used to characterize protein abundance changes during an EMT event. The LC-MS/MS analysis and label-free quantification revealed that TGF beta and K-Ras(v12) induce a similar pattern of protein regulation and that besides the expected cytoskeletal changes, a strong increase in the anabolism and energy production machinery was observed.Biological SignificanceTo our knowledge, this is the first proteomic analysis combining a label-free quantification with an SRM validation of proteins regulated by TGF beta and K-Ras(v12). This study reveals new insights in the characterization of the changes occurring during an epithelial-mesenchymal transition (EMT) event. Notably, a strong increase in the anabolism and energy production machinery was observed upon both EMT inducers

Interaction of the Epstein-Barr Virus mRNA Export Factor EB2 with Human Spen Proteins SHARP, OTT1, and a Novel Member of the Family, OTT3, Links Spen Proteins with Splicing Regulation and mRNA Export

International audienceThe Epstein-Barr virus early protein EB2 (also called BMLF1, Mta, or SM), a protein absolutely required for the production of infectious virions, shares properties with mRNA export factors. By using a yeast two-hybrid screen, we have identified the human protein OTT3 as an EB2-interacting factor. OTT3 is a new member of the Spen (split end) family of proteins (huSHARP, huOTT1, DmSpen, and muMINT), which are characterized by several N-terminal RNA recognition motifs and a highly conserved C-terminal SPOC (Spen Paralog and Ortholog C-terminal) domain that, in the case of SHARP, has been shown to interact with SMRT/NCoR co-repressors. OTT3 is ubiquitously expressed as a 120-kDa protein. Transfected OTT3 is a nonshuttling nuclear protein that co-localizes with co-transfected EB2. We also showed that EB2 interacts with the SPOC domains of both OTT1 and SHARP proteins. Although the OTT3 interaction domain maps within the 40 N-terminal amino acids of EB2, OTT1 and SHARP interact within the C-terminal half of the protein. Furthermore, we demonstrated that the capacity of the OTT3 and OTT1 SPOC domains to interact with SMRT and repress transcription is far weaker than that of SHARP. Thus there is no evidence for a role of OTT3 in transcriptional regulation. Most interestingly, however, we have found that OTT3 has a role in splicing regulation; OTT3 represses accumulation of the alternatively spliced-thalassemia mRNAs, but it has no effect on the-globin constitutively spliced mRNA. Thus our results suggested a new function for Spen proteins related to mRNA export and splicin