Molecular basis of LH action on breast cancer cell migration and invasion via kinase and scaffold proteins

Breast cancer (BC) is a major public health problem affecting women worldwide. Approximately 80% of diagnosed cases are hormone-dependent breast cancers. These hormones are known to stimulate tumor development and progression. In this setting, tentative evidence suggests that luteinizing hormone (LH) may also play a role in tumors. In BC cells that express functional LH receptors (LHR), this hormone regulates cell migration and invasion by controlling several kinases that activate actin cytoskeletal proteins. In this article, we show that LH induces phosphorylation of paxillin and its translocation toward the plasmatic membrane, where focal adhesion complexes are assembled. This process is triggered via a rapid extra-gonadal LHR signaling to Src/FAK/paxillin, which results in the phosphorylation/activation of the nucleation promoter factors cortactin and N-WASP. As a consequence, Arp2/3 complexes induce actin polymerization, essential to promote cell adhesion, migration, and invasion, thus enhancing metastatic spread of tumoral cells. Our findings provide relevant information about how gonadotrophins exert their action in BC. This information helps us understand the extragonadal effects of LH on BC metastasis. It may provide new perspectives for therapeutic treatment, especially for women with high serum levels of gonadotrophins.Fil: Mondaca, Joselina Magali. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Uzair, Ivonne Denise. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Castro Guijarro, Ana Carla. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Flamini, Marina Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Sanchez, Angel Matias. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; Argentin

A decade of user operation on the macromolecular crystallography MAD beamline ID14-4 at the ESRF

The improvement of the X-ray beam quality achieved on ID14-4 by the installation of new X-ray optical elements is described

Experimental procedure for the characterization of radiation damage in macromolecular crystals

A novel automatic procedure to determine the sensitivity of macromolecular crystals to radiation damage is presented. The information extracted from this procedure can be directly used for optimal planning of data collection or/and beamline calibration

The ID23-2 structural biology microfocus beamline at the ESRF

Beamline ID23-2, the first dedicated and highly automated high-throughput monochromatic macromolecular crystallography microfocus beamline, is described

MxCuBE: a synchrotron beamline control environment customized for macromolecular crystallography experiments

MxCuBE is a beamline control environment optimized for the needs of macromolecular crystallography. This paper describes the design of the software and the features that MxCuBE currently provides

Heregulin-induced cell migration is prevented by trastuzumab and trastuzumab-emtansine in HER2+ breast cancer

Purpose: Heregulin (HRG) signaling has been implicated in the development of an aggressive phenotype in breast cancer (BC) cells, and HER2 overexpression has been associated with a worse prognosis in BC patients. Nevertheless, the molecular mechanisms through which HRG affects the efficiency of anti-HER2 therapies such as trastuzumab (Tz) and trastuzumabemtansine (T-DM1) are currently unknown. Methods: In the present study, we evaluate the molecular action of HRG toward fundamental scaffold proteins and several kinases in the signal transduction pathways triggered via HER2/HER3, which integrate precise and sequential steps to promote changes in cell morphology to impulse BC cell migration. In addition, we evaluate the effectiveness of Tz and T-DM1 on the control of key proteins involved in BC cell motility, since the acquisition of a migratory phenotype is essential to promote invasion and metastasis. Results: We show that HRG induces actin cytoskeleton reorganization and focal adhesion complex formation, and promotes actin nucleation in BT-474 BC cells. This signaling is triggered by HER2/HER3 to c-Src, FAK and paxillin. When paxillinis phosphorylated, it recruits PAK1, which then phosphorylates cortactin. In parallel, paxillin signals to N-WASP, and both signalings regulate Arp2/3 complex, leading to the local reorganization of actin fibers. Conclusions: Our findings reveal an original mechanism by which HRG increases HER2+ BC cell motility, and show that the latter can be abolished by Tz and T-DM1 treatments. These results provide evidence for the molecular mechanisms involved in cell motility and drug resistance. They will be useful to develop new and more specific therapeutic schemes that interfere with the progression and metastasis of HER2+ BC.Fil: Mondaca, Joselina Magali. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Castro Guijarro, Ana Carla. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Flamini, Marina Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Sanchez, Angel Matias. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; Argentin

MXCuBE3 Bringing MX Experiments to the WEB

International audienceOriginally conceived at ESRF and first deployed in 2005 MXCuBE, Macromolecular Xtallography Customized Beamline Environment, has with its successor MXCuBE2, become a successful international collaboration. The aim of the collaboration is to develop a beamline control application for macromolecular crystallography (MX) that are independent of underlying instrument control software and thus deployable at the MX beamlines of any synchrotron source. The continued evolution of the functionality offered at MX beamlines is to a large extent facilitated by active software development. New demands and advances in technology have led to the development of a new version of MXCuBE, MXCuBE3, The design of which was inspired by the results of a technical pre-study and user survey. MXCuBE3 takes advantage of the recent development in web technologies such as React and Redux to create an intuitive and user friendly application. The access to the application from any web browser further simplifies the operation and natively facilitates the execution of remote experiments

BLISS - Experiments Control for ESRF EBS Beamlines

International audienceBLISS is the new ESRF control system for running experiments, with full deployment aimed for the end of the EBS upgrade program in 2020. BLISS provides a global approach to run synchrotron experiments, thanks to hardware integration, Python sequences and an advanced scanning engine. As a Python package, BLISS can be easily embedded into any Python application and data management features enable online data analysis. In addition, BLISS ships with tools to enhance scientists user experience and can easily be integrated into TANGO based environments, with generic TANGO servers on top of BLISS controllers. BLISS configuration facility can be used as an alternative TANGO database. Delineating all aspects of the BLISS project from beamline device configuration up to the integrated user interface, this talk will present the technical choices that drove BLISS design and will describe the BLISS software architecture and technology stack in depth

BLISS - Experiments Control for ESRF Beamline

International audienceBLISS is the new ESRF control system for running experiments, with full deployment aimed for the end of the EBS upgrade program in 2020. BLISS provides a global approach to run synchrotron experiments, thanks to hardware integration, Python sequences and an advanced scanning engine. As a Python package, BLISS can be easily embedded into any Python application and data management features enable online data analysis. In addition, BLISS ships with tools to enhance scientists user experience and can easily be integrated into TANGO based environments, with generic TANGO servers on top of BLISS controllers. BLISS configuration facility can be used as an alternative TANGO database. Delineating all aspects of the BLISS project from beamline device configuration up to the integrated user interface, this poster will present the technical choices that drove BLISS design and will describe the BLISS software architecture and technology stack in depth

Combination Treatment of Retinoic Acid Plus Focal Adhesion Kinase Inhibitor Prevents Tumor Growth and Breast Cancer Cell Metastasis

All-trans retinoic acid (RA), the primary metabolite of vitamin A, controls the development and homeostasis of organisms and tissues. RA and its natural and synthetic derivatives, both known as retinoids, are promising agents in treating and chemopreventing different neoplasias, including breast cancer (BC). Focal adhesion kinase (FAK) is a crucial regulator of cell migration, and its overexpression is associated with tumor metastatic behavior. Thus, pharmaceutical FAK inhibitors (FAKi) have been developed to counter its action. In this work, we hypothesize that the RA plus FAKi (RA + FAKi) approach could improve the inhibition of tumor progression. By in silico analysis and its subsequent validation by qPCR, we confirmed RARA, SRC, and PTK2 (encoding RARα, Src, and FAK, respectively) overexpression in all breast cells tested. We also showed a different pattern of genes up/down-regulated between RA-resistant and RA-sensitive BC cells. In addition, we demonstrated that both RA-resistant BC cells (MDA-MB-231 and MDA-MB-468) display the same behavior after RA treatment, modulating the expression of genes involved in Src-FAK signaling. Furthermore, we demonstrated that although RA and FAKi administered separately decrease viability, adhesion, and migration in mammary adenocarcinoma LM3 cells, their combination exerts a higher effect. Additionally, we show that both drugs individually, as well as in combination, induce the expression of apoptosis markers such as active-caspase-3 and cleaved-PARP1. We also provided evidence that RA effects are extrapolated to other cancer cells, including T-47D BC and the human cervical carcinoma HeLa cells. In an orthotopic assay of LM3 tumor growth, whereas RA and FAKi administered separately reduced tumor growth, the combined treatment induced a more potent inhibition increasing mice survival. Moreover, in an experimental metastatic assay, RA significantly reduced metastatic lung dissemination of LM3 cells. Overall, these results indicate that RA resistance could reflect deregulation of most RA-target genes, including genes encoding components of the Src-FAK pathway. Our study demonstrates that RA plays an essential role in disrupting BC tumor growth and metastatic dissemination in vitro and in vivo by controlling FAK expression and localization. RA plus FAKi exacerbate these effects, thus suggesting that the sensitivity to RA therapies could be increased with FAKi coadministration in BC tumors