Laboratory of Genitourinary Cancer Pathogenesis Microscopy Core Facility

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The Core Facility houses a Zeiss LSM 510 META scanning module mounted on an Axio Observer Z1 motorized fluorescent microscope, an upright Axioplan 2 fluorescent microscope equipped with high resolution lenses and Nomarski DIC optics, and an Axio Observer Z1 fluorescent microscope, computer and monitor equipped with axiovision software for image acquisition. In addition, The facility houses a VivaView fluorescent incubator microscope capable of imaging cells growing in culture over several days or weeks. Researchers from fifteen different laboratories (approximately 40 users) in the NCI are currently using the Confocal Core Facility. The research focus of the scientists using the facility includes ovarian, breast, prostate, thyroid, and lung cancer progression and metastasis, chemotherapy used in pediatric oncology, and AIDS therapy. In the Cell and Cancer Biology Branch (CCBB), 21 scientists including staff scientists, postdoctoral fellows, post baccalaureate fellows, and summer students routinely use the core facility in their research efforts. Postdoctoral fellows in Dr. Zheng-Gang Lius laboratory are using nuclear microinjection of DNA to investigate the role of death domain proteins in apoptosis. Confocal microscopy is being used by this group to investigate the molecular mechanisms of endoplasmic reticulum stress and cellular localization of proteins involved in apoptosis. In Dr. Kathy Kelly's laboratory three postdoctoral fellows and one post baccalaureate fellow are using confocal microscopy to study molecular mechanisms and identify differentiation markers and signaling molecules on primary murine prostate cells in order to elucidate the signaling pathways involved in prostate cancer metastasis. They are also characterizing normal and transformed prostate epithelial cells stained with progression markers and signaling molecules in 2 and 3 dimensional cultures. Confocal microscopy is essential for observing the localization of these components in the spherical and tubular structures that grow in 3D culture. A research fellow and a post baccalaureate fellow are using confocal microscopy for imaging brain metastasis from fluorescently labeled prostate cancer cells. Scientists from several other branches or laboratories are currently using the core as an integral part of their research. Below are some examples of ongoing projects using the core facility. 1.Lab of Cancer Biology and Genetics: Study of nucleoside analogs used in AIDS therapy. 2.Mammary Biology and Tumorigenesis: Investigation of the mammary stem cell niche and its ability to reprogram progenitor populations to the embryonic stem (ES) cell. Confocal microscopy is being used to confirm the presence of the Y-chromosome in the ES-derived cells and to establish that these cells are functional in their current location by milk protein immunohistochemistry. 3.Medical Oncology Branch: a.The role of the GXXXG motif in the homodimerization of the ABC half-transporter, ABCG2, which confers resistance to several important clinical agents b.Localization of p53 in cancer cell lines and colocalization of p53 with tubulin and importin c.Effect of platinum compounds on the microtubule cytoskeleton. These are essential studies that will be correlated to clinical trial data and information from these studies may prove to be important in the development of therapies to alleviate the toxicity associated with these compounds. Apart from collaborating on projects involving the confocal microscopy core facility, teaching scientists various aspects of confocal microscopy and microinjection, and maintaining the equipment in the core, I am conducting research on the mechanisms of CD97 signal transduction and the role of CD97 in cancer progression and metastasis in collaboration with the branch Chief of CCBB, Kathy Kelly. The details of this project are described in Kathy Kelly's annual report. CD97, an adhesion class G protein coupled receptor, is expressed on inflammatory cells and several carcinomas. CD97 expression increases in parallel with malignant grade in thyroid, esophageal, gastric, colorectal, and prostate tumors. We have demonstrated that CD97 acts both as a cell-autonomous receptor on tumor cells and as a ligand for integrins alpha5 beta1 and alphav beta3 on endothelial cells. We showed that in its capacity as a receptor, CD97 signaling couples through the G-alpha 12/13 family of heterotrimeric G proteins resulting in increased Rho activity. An analysis of prostate and thyroid tumor cell lines has shown that abnormal overexpression of endogenously-expressed CD97 seems to lead to ligand-independent signaling. Depletion of endogenous CD97 in prostate tumor cell lines resulted in decreased metastasis to bone. We have two recent publications demonstarting that CD97 plays an active role in prostate and thyroid cancer progression probably via heterodimerization with the lysophosphatidic receptor.