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3. Global genome analysis reveals a vast and dynamic anellovirus landscape within the human virome

Author

Arze, Cesar A., primary, Springer, Simeon, additional, Dudas, Gytis, additional, Patel, Sneha, additional, Bhattacharyya, Agamoni, additional, Swaminathan, Harish, additional, Brugnara, Carlo, additional, Delagrave, Simon, additional, Ong, Tuyen, additional, Kahvejian, Avak, additional, Echelard, Yann, additional, Weinstein, Erica G., additional, Hajjar, Roger J., additional, Andersen, Kristian G., additional, and Yozwiak, Nathan L., additional

Published
2021
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5. List of Contributors

Author

Alicea, Bradly, primary, Amano, Tomokazu, additional, Ambroggio, Jennifer, additional, Amarnath, Dasari, additional, Campbell, Keith H.S., additional, Surani, M. Azim, additional, Baeumer, Nicole, additional, Balbach, Sebastian T., additional, Bertolini, Marcelo, additional, Beyhan, Zeki, additional, Boiani, Michele, additional, Byrne, James A., additional, Callesen, Henrik, additional, Canovas, Sebastian, additional, Chavatte-Palmer, Pascale, additional, Chen, LiHow, additional, Choi, Inchul, additional, Cibelli, José B., additional, Colleoni, Silvia, additional, Crosetto, Nicola, additional, Dinnyes, Andras, additional, Drexler, Hannes C.A., additional, Duchi, Roberto, additional, Echelard, Yann, additional, Egli, Dieter, additional, Faber, David, additional, Fan, Jason, additional, First, Neal L., additional, Fissore, Rafael A., additional, Fuellen, Georg, additional, Fulka, Josef, additional, Galli, Cesare, additional, Gardner, David K., additional, Gavin, William, additional, Green, Ronald M., additional, Gurdon, J.B., additional, Hirano, Kunio, additional, Hoshino, Yoichiro, additional, Howard, Harlan, additional, Inoue, Kimiko, additional, Ishino, Fumitoshi, additional, Ito, Junya, additional, Jammes, Hélène, additional, Jones, Jeff, additional, Jones, Kathleen M., additional, Kent-First, Marijo, additional, Kato, Yoko, additional, Kishigami, Satoshi, additional, Ko, Minoru S.H., additional, Kohda, Takashi, additional, Lagutina, Irina, additional, Lane, Michelle, additional, Latham, Keith E., additional, Lazzari, Giovanna, additional, Lee, Byeong Chun, additional, Lee, Jiyoung, additional, Lee, Kiho, additional, Lee, Rita, additional, Loi, Pasqualino, additional, Malcuit, Christopher M., additional, Masiello, Nick, additional, Meade, Harry, additional, Meng, Qinggang, additional, Mitalipov, Shoukhrat, additional, Mizutani, Eiji, additional, Modlinski, Jacek, additional, Nguyen, Van Thuan, additional, Niemann, Heiner, additional, Ogura, Atsuo, additional, Page, Raymond L., additional, Paudel, Yogesh, additional, Paull, Daniel, additional, Pfeiffer, Martin J., additional, Piedrahita, Jorge A., additional, Pinmee, Boonya, additional, Polgar, Zsuzsanna, additional, Pommer, Jerry, additional, Prather, Randall S., additional, Pruett-Miller, Shondra M., additional, Ptak, Grazyna, additional, Rudenko, Larisa, additional, Saeki, Kazuhiro, additional, Schatten, Gerald, additional, Schmidt, Mette, additional, Schofield, Michael, additional, Seidel, George E., additional, Siatkowski, Marcin, additional, Simerly, Calvin, additional, Siripattarapravat, Kannika, additional, St. John, Justin C., additional, Stice, Steven L., additional, Sullivan, Eddie J., additional, Tachibana, Masahito, additional, Tada, Takashi, additional, Tancos, Zsuzsanna, additional, Taniguchi, Shunji, additional, Teperek-Tkacz, Marta, additional, Tian, Xiuchun (Cindy), additional, Trounson, Alan, additional, Tso Sun, Liang, additional, Tsunoda, Yukio, additional, Wakai, Takuya, additional, Wakamatsu, Yuko, additional, Wakayama, Sayaka, additional, Wakayama, Teruhiko, additional, Wang, Zhongde, additional, Zhang, Xia, additional, and Zhu, Jie, additional

Published
2014
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8. Viable Transgenic Goats Derived from Skin Cells

Author

Behboodi, Esmail, Memili, Erdogan, Melican, David T., Destrempes, Margaret M., Overton, Susan A., Williams, Jennifer L., Flanagan, Peter A., Butler, Robin E., Liem, Hetty, Chen, Li How, Meade, Harry M., Gavin, William G., and Echelard, Yann

Published
2004
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11. List of Contributors

Author

Ahlstrom, Jon D., primary, Ahsan, Taby, additional, Allickson, Julie, additional, Almarza, Alejandro J., additional, Anderson, James M., additional, Andrews, Peter, additional, Aslan, Hadi, additional, Atala, Anthony, additional, Badylak, Stephen F., additional, Batra, Ashok, additional, Baumann, M. Douglas, additional, Bellamkonda, Ravi V., additional, Bergman, Nicole M., additional, Beyhan, Z., additional, Bhatia, Mickie, additional, Bhatia, Sangeeta N., additional, Black, Peter M., additional, Blau, Helen, additional, Boden, Scott D., additional, Brey, Eric M., additional, Brivanlou, Ali H., additional, Brown, Chris R., additional, Bruder, Scott P., additional, Chambers, S.M., additional, Chen, Christopher S., additional, Chen, LiHow, additional, Chen, Mike, additional, Chen, Sulin, additional, Cheng, N., additional, Christ, George J., additional, Chung, Seyung, additional, Cibelli, J.B., additional, Cimini, Massimo, additional, De Coppi, Paolo, additional, Dodla, Mahesh C., additional, Dominguez-Bendala, Juan, additional, Doyle, AM, additional, Durfor, Charles N., additional, Echelard, Yann, additional, Effros, Rita B., additional, Elisseeff, Jennifer, additional, Ellis, Ewa C.S., additional, Emamaullee, Juliet A., additional, Erickson, Carol A., additional, De Filippo, Roger, additional, Fink, Donald, additional, Fissell, William H., additional, Friedlaender, Gary E., additional, Furth, Mark E., additional, Gafni, Yossi, additional, Gage, Keneth, additional, Garcia, Andres, additional, Gavin, William, additional, Gazit, Daniel, additional, Gazit, Zulma, additional, Gemmiti, Christopher S., additional, Gerlach, Jörg C., additional, Gokhale, Paul J., additional, Goodell, M.A., additional, Griffith, May, additional, Guenin, Louis M., additional, Giuliani, Stefano, additional, Guldberg, Robert E., additional, Hacker, M.C., additional, Harrison, Benjamin S., additional, Hartmann, Bernd, additional, Hilbert, Stephen H., additional, Hillel, Alexander, additional, Hipp, Jason, additional, Holcomb, Col. J.B., additional, Hollinger, Jeffrey O., additional, Holy, Chantal E., additional, Hout, Mariah, additional, Hu, Jiang, additional, Huang, George T.-J., additional, Huard, Johnny, additional, Hui, Elliot E., additional, Humes, H. David, additional, Intaglietta, Marcos, additional, Ji, Junfeng, additional, Jiang, Yueha, additional, Johnen, Christa, additional, Johnston, Josephine, additional, Joraku, Akira, additional, Kaplan, David L., additional, Kaplan, David S., additional, Khang, Gilson, additional, Khanzada, Rehan N., additional, Kim, Soon Hee, additional, Kim, Moon Suk, additional, Kimelman, Nadav, additional, Klimanskaya, Irina, additional, Kluge, Jonathan A., additional, Kolambkar, Yash, additional, Koh, Chester J., additional, Komura, Makoto, additional, Kondziolka, Douglas, additional, Konya, Deniz, additional, Kues, Wilfried A., additional, Kwong, Francois Ng kee, additional, Lamba, Deepak, additional, Lee, Hai Bang, additional, Lee, Hyukjin, additional, Leisk, Gary G., additional, Leong, Kam W., additional, Levine, Ariel J., additional, Li, Ren Ke, additional, Li, Wan-Ju, additional, Lim, Grace J., additional, Lin, Yan, additional, Lindblad, William J., additional, Liu, Wendy F., additional, Liu, Xiaohua, additional, Lucas-Hahn, Andrea, additional, Luttun, Aernout, additional, Lynch, Samuel, additional, Ma, Peter X., additional, Maher, Ellen, additional, Martins-Green, Manuela, additional, McClelland, Randall E., additional, McFarland, Richard, additional, McIntire, Larry V., additional, Meade, Harry, additional, Melican, David L., additional, Mikos, A.G., additional, Montoya, Fernando Ulloa, additional, Nerem, Robert M., additional, Niemann, Heiner, additional, Nori, Aparna, additional, Pabst, Patrea L., additional, Park, Kook In, additional, Park, Tae Gwan, additional, Patterson, David P., additional, Pauwelyn, Karen, additional, Pelled, Gadi, additional, Perin, Laura, additional, Petreaca, M., additional, Pileggi, Antonello, additional, Pomerantz, Jason H., additional, Porter, Blaise, additional, Radisic, Milica, additional, Ratner, Buddy D., additional, Reddi, A. Hari, additional, Reh, Thomas A., additional, Reid, Lola M., additional, Ricordi, Camillo, additional, Ross, Jeff, additional, Russell, Alan J., additional, Santos, Filipe N.C., additional, Schmitz, John P., additional, Schuch, Gunter, additional, Sedrakyan, Sargis, additional, Sefton, Michael V., additional, Serafini, Marta, additional, Shah, Paulesh, additional, Shapiro, A.M. James, additional, Sheardown, Heather, additional, Shoichet, Molly S., additional, Siddiqui, M. Minhaj, additional, Sidman, Richard L., additional, Silverman, Ronald, additional, Skuk, Daniel, additional, Snyder, Evan, additional, Soker, Shay, additional, Spector, Myron, additional, Stocum, David L., additional, Strom, Stephen C., additional, Taylor, Doris A., additional, Teng, Yang D., additional, Thomson, James, additional, Tranquillo, Robert T., additional, Tremblay, Jacques P., additional, Tsai, Amy, additional, Tuan, Rocky S., additional, Tubo, Ross S., additional, Van Dyke, Mark, additional, Vavoie, Deborah, additional, Verfaillie, Catherine, additional, Volenec, F. Jerry, additional, Wargo, Sara, additional, Warnwath, Joseph W., additional, Wechsler, Lawrence, additional, Wei, Shen, additional, Weisel, Richard D., additional, West, Jennifer L., additional, Williams, Chrysanthi, additional, Williams, J. Koudy, additional, Witten, Celia, additional, Wolf, Steven E., additional, Woo, Savio L.-Y., additional, Wosnick, Jordan H., additional, Wrenzycki, Christine, additional, Xaymardan, Munira, additional, Yao, Hsin-Lei, additional, Yazdani, Saami K., additional, Yelick, Pamela C., additional, Yoo, James J., additional, Yu, Junying, additional, Zeilinger, Katrin, additional, Zeng, Lepeng, additional, Zenovich, Andrey G., additional, Zhong, Bonan, additional, Ziomek, Carol A., additional, and Zoloth, Laurie, additional

Published
2008
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12. The neuronal repressor REST/NRSF is an essential regulator in medulloblastoma cells

Author

Lawinger, Patrick, Venugopal, Radjendirane, Guo, Zong-Sheng, Immaneni, Anand, Sengupta, Devjani, Lu, Wenying, Rastelli, Luca, Marin Dias Carneiro, Ana, Levin, Victor, Fuller, Gregory N., Echelard, Yann, and Majumder, Sadhan

Subjects
Cancer cells -- Physiological aspects, Cancer cells -- Genetic aspects, Cancer cells -- Research, DNA binding proteins -- Physiological aspects, DNA binding proteins -- Research, Medulloblastoma -- Research, Genes -- Physiological aspects, Genes -- Research
Abstract

Medulloblastoma is the most malignant pediatric brain tumor. It is believed to originate from the undifferentiated external granule layer cells in the cerebellum, but the mechanism of tumorigenesis remains unknown. Here we studied three types of human medulloblastoma cells that express markers corresponding to different levels of neuronal differentiation. They expressed the neuronal repressor element 1 (RE1) silencing transcription factor/neuron-restrictive silencer factor (REST/NRSF; refs. 7-10) at very high levels compared with either neuronal progenitor NTera2 (NT2) cells or fully differentiated human neuron teratocarcinoma (hNT cells). To counter the effect of REST/NRSF, we used a recombinant transcription factor, REST-VP16, constructed by replacing repressor domains of REST/NRSF with the activation domain of viral protein (VP16). Transient expression of REST-VP16 in medulloblastoma cells was able to compete with the endogenous REST/NRSF for DNA binding and stimulate neuronal promoters. High-efficiency expression of REST-VP16 mediated by adenovirus vectors (Ad.REST-VP16) in medulloblastoma cells was able to counter REST/NRSF-mediated repression of neuronal promoters, stimulate expression of endogenous neuronal genes and trigger apoptosis through the activation of caspase cascades. Furthermore, intratumoral injection of Ad.REST-VP16 in established medulloblastoma tumors in nude mice inhibited their growth. Therefore, REST/NRSF may serve as a new target for therapeutic interventions for medulloblastoma through agents such as REST-VP16., Author(s): Patrick Lawinger [1, 4]; Radjendirane Venugopal [1, 4]; Zong-Sheng Guo [2, 4]; Anand Immaneni [1]; Devjani Sengupta [1]; Wenying Lu [1]; Luca Rastelli [1]; Ana Marin Dias Carneiro [1]; [...]

Published
2000
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17. A 5.5-kb enhancer is both necessary and sufficient for regulation of Wnt-1 transcription in vivo

Author

Danielian, Paul S., Echelard, Yann, Vassileva, Galya, and McMahon, Andrew P.

Subjects
Central nervous system -- Research, Genetically modified mice -- Analysis, Mesencephalon -- Research, Embryology -- Research, Biological sciences
Abstract

Writ-1 encodes a secreted signaling molecule which is required for development of the midbrain and anterior hindbrain in the mouse. Wnt-1 expression is initiated early in the development of the central nervous system in a region predicted to give rise to the midbrain. Later in development Wnt-1 expression is restricted to regions of the forebrain, midbrain, and spinal cord. Previous studies identified a 5.5-kb enhancer in the Wnt-1 locus which is sufficient to activate transcription of a reporter gene in a pattern very similar to that of the endogenous Wnt-1 gene. Here we have assessed if this enhancer is an important component of the endogenous regulatory sequences of Wnt-1 by gene targeting and by testing if it is able to express Wnt-1 in a pattern which is sufficient to rescue the phenotype of loss of Wnt-1. Our results show that the 5.5-kb enhancer is both necessary and sufficient for Wnt-1 expression in vivo.

Published
1997

18. Production of recombinant therapeutic proteins in the milk of transgenic animals

Author

Echelard, Yann, Ziomek, Carol A., and Meade, Harry M.

Subjects
Biopolymers, Erythropoietin, Fibrin, Biopharmaceutics, Medical research, Medicine, Experimental, Genetic research, Gamma globulins, Gene expression, Antithrombin III, Dairy cattle, Drug approval, Genetic engineering, Genetic disorders, Animal feeding and feeds, Dairy farming, Galactose, Proteins, Blood coagulation factors, Biotechnology industry, Business, Pharmaceuticals and cosmetics industries
Abstract

ABSTRACT Transgenic milk production offers a cost-effective system for the manufacturing of large amounts of complex proteins. Specifically, commercialization is near for recombinant human antithrombin (rhAT) expressed in transgenic dairy [...]

Published
2006

19. Sonic hedgehob, a member of a family of putative signaling molecules, is implicated in the regulation of CNS polarity

Author

Echelard, Yann, Epstein, Douglas J., St-Jacques, Benoit, Liya Shen, Mohler, Jym, McMahon, Jill A., and McMahon, Andrew P.

Subjects
Central nervous system -- Physiological aspects, Drosophila -- Research, Developmental biology -- Research, Mice -- Research, Biological sciences
Abstract

The Sonic hedgehog (Shh) gene is expressed in the areas that mediate central nervous system (CNS) and limb polarity, indicating its role in the inductive interactions that form the CNS. The Shh and two other closely related genes encode putative secreted proteins and are possibly involved in cell-cell interactions. Ectopic expression of Shh in the mouse CNS triggers the flow of plate-expressed genes.

Published
1993

22. Ectopic Expression of Olig1 Is Deleterious to V3 Interneuron Development in the Developing CNS

Author

Sun, Tao, Lu, Richard, Stiles, Charles, Echelard, Yann, and Rowitch, David

Subjects
Developmental neurology -- Genetic aspects, Neural tube -- Research, Biological sciences
Abstract

In the developing neural tube, precursors at defined positions along the dorsal-ventral axis respond to Sonic hedgehog (Shh) secreted from the notocord and floorplate and elaborate distinct neurons identified by homeodomain protein expression. Oligodendrocyte lineage genes (Olig1/2) that encode a novel class of bHLH proteins are expressed in the oligodendrocyte lineage in the CNS. Moreover, their early expression is regulated by Shh in a dosage-dependent manner, suggesting that Olig genes may play roles in pattern formation and neuronal and/or glial cell fate determination. To study possible Olig functions during development, we analyzed endogenous Olig1/2 expression and found that Olig genes were initially expressed in the mouse spinal cord as early as 8.5 dpc and that expression at 11.5 dpc occurred in a restricted domain. Olig expression overlaps Nkx2.2 expression, where V3 interneurons (INs) are generated. We next generated transgenic mice that ectopically express Olig1 in the ventricular zone under control of the Hoxb4 enhancer. Global disruption of pattern was not observed by analysis of markers for neural precursors (NPs), motor neurons, and V2 INs. In contrast, misexpression Oligl evidently is detrimental to production of V3 INs, as Sim1-expressing cells were absent in these transgenic mice. Moreover, increased levels of cell death were observed. These data suggest that Olig genes may antagonize the development of certain neuronal subtypes and may refine pattern formation in NPs of the ventral neural tube.

Published
2001

27. TG20, a Transgenically-Derived Anti-CD20 Monoclonal Antibody, Exhibits Enhanced Cytotoxicity Against Cells with Low Levels of CD20

Author

Echelard, Yann, primary, Pollock, Daniel P., additional, Coupade, Catherine De, additional, Olivier, Aurélie Groseil, additional, Brune, Frédérique, additional, Chen, LiHow, additional, Masiello, Nicholas C., additional, Williams, Jennifer L., additional, Gavin, William G., additional, Chtourou, Sami, additional, and Meade, Harry M., additional

Published
2011
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28. Adeno‐associated virus (AAV)‐mediated transduction of male germ line stem cells results in transgene transmission after germ cell transplantation

Author

Honaramooz, Ali, primary, Megee, Susan, additional, Zeng, Wenxian, additional, Destrempes, Margret M., additional, Overton, Susan A., additional, Luo, Jinping, additional, Galantino-Homer, Hannah, additional, Modelski, Mark, additional, Chen, Fangping, additional, Blash, Stephen, additional, Melican, David T., additional, Gavin, William G., additional, Ayres, Sandra, additional, Yang, Fang, additional, Wang, P. Jeremy, additional, Echelard, Yann, additional, and Dobrinski, Ina, additional

Published
2007
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31. Effects of Long-term Administration of High-dose Recombinant Human Antithrombin in Immunosuppressed Primate Recipients of Porcine Xenografts

Author

Cozzi, Emanuele, primary, Simioni, Paolo, additional, Boldrin, Massimo, additional, Seveso, Michela, additional, Calabrese, Fiorella, additional, Baldan, Nicola, additional, Busetto, Roberto, additional, Tormene, Daniela, additional, Gavasso, Sabrina, additional, Castagnaro, Massimo, additional, Echelard, Yann, additional, Rice, Tim, additional, Plebani, Mario, additional, Carraro, Paolo, additional, Bosio, Erika, additional, Valente, Marialuisa, additional, Pagnan, Antonio, additional, Thiene, Gaetano, additional, and Ancona, Ermanno, additional

Published
2005
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40. Production of goats by somatic cell nuclear transfer

Author

Baguisi, Alexander, primary, Behboodi, Esmail, additional, Melican, David T., additional, Pollock, Julie S., additional, Destrempes, Margaret M., additional, Cammuso, Christine, additional, Williams, Jennifer L., additional, Nims, Scott D., additional, Porter, Catherine A., additional, Midura, Patricia, additional, Palacios, Monica J., additional, Ayres, Sandra L., additional, Denniston, Richard S., additional, Hayes, Michael L., additional, Ziomek, Carol A., additional, Meade, Harry M., additional, Godke, Robert A., additional, Gavin, William G., additional, Overström, Eric W., additional, and Echelard, Yann, additional

Published
1999
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45. Adeno-associated virus (AAV)-mediated transduction of male germ line stem cells results in transgene transmission after germ cell transplantation.

Author

Honaramooz, Ali, Megee, Susan, Wenxian Zeng, Destrempes, Margret M., Overton, Susan A., Jinping Luo, Galantino-Homer, Hannah, Modelski, Mark, Fangping Chen, Blash, Stephen, Melican, David T., Gavin, William G., Ayres, Sandra, Fang Yang, Wang, P. Jeremy, Echelard, Yann, and Dobrinski, Ina

Subjects
STEM cells, GERM cells, CELL transplantation, VIRUSES, GENETIC vectors
Abstract

We explored whether exposure of mammalian germ line stem cells to adeno-associated virus (AAV), a gene therapy vector, would lead to stable transduction and transgene transmission. Mouse germ cells harvested from experimentally induced crypt-orchid donor testes were exposed in vitro to AAV vectors carrying a GFP transgene and transplanted to germ cell-depleted syngeneic recipient testes, resulting in colonization of the recipient testes by transgenic donor cells. Mating of recipient males to wild-type females yielded 10% transgenic offspring. To broaden the approach to nonrodent species, AAV-transduced germ cells from goats were transplanted to recipient males in which endogenous germ cells had been depleted by fractionated testicular irradiation. Transgenic germ cells colonized recipient testes and produced transgenic sperm. When semen was used for in vitro fertilization (IVF), 10% of embryos were transgenic. Here, we report for the first time that AAV-mediated transduction of mammalian germ cells leads to transmission of the transgene through the male germ line. Equally important, this is also the first report of transgenesis via germ cell transplantation in a nonrodent species, a promising approach to generate transgenic large animal models for biomedical research. [ABSTRACT FROM AUTHOR]

Published
2008
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47. Fertility and Germline Transmission of Donor Haplotype Following Germ Cell Transplantation in Immunocompetent Goats1

Author

Honaramooz, Ali, Behboodi, Esmail, Megee, Susan O., Overton, Susan A., Galantino-Homer, Hannah, Echelard, Yann, and Dobrinski, Ina

Abstract

Transplantation of spermatogonial stem cells into syngeneic or immunosuppressed recipient mice or rats can result in donor-derived spermatogenesis and fertility. Recently, this approach has been employed to introduce a transgene into the male germline. Germ-cell transplantation in species other than laboratory rodents, if successful, holds great promise as an alternative to the inefficient methods currently available to generate transgenic farm animals that can produce therapeutic proteins in their milk or provide organs for transplantation to humans. To explore whether germ-cell transplantation could result in donor-derived spermatogenesis and fertility in immunocompetent recipient goats, testis cells were transplanted from transgenic donor goats carrying a human alpha-1 antitrypsin expression construct to the testes of sexually immature wild-type recipient goats. After puberty, sperm carrying the donor-derived transgene were detected in the ejaculates of two out of five recipients. Mating of one recipient resulted in 15 offspring, one of which was transgenic for the donor-derived transgene. This is the first report of donor cell-derived sperm production and transmission of the donor haplotype to the next generation after germ-cell transplantation in a nonrodent species. Furthermore, these results indicate that successful germ-cell transplantation is feasible between immunocompetent, unrelated animals. In the future, transplantation of genetically modified germ cells may provide a more efficient alternative for production of transgenic domestic animals.

Published
2003
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48. Efficiency of Cationic Lipid-Mediated Transfection of Polarized and Differentiated Airway Epithelial Cells In Vitroand In Vivo

Author

Jiang, Canwen, O'Connor, Sean P., Fang, Shaona L., Wang, Kathryn X., Marshall, John, Williams, Jennifer L., Wilburn, Brian, Echelard, Yann, and Cheng, Seng H.

Abstract

ABSTRACTSystematic analysis of a large number of different cationic lipids has led to the identification of novel structures (GL-67) and formulations of cationic lipid:plasmid DNA (pDNA) complexes that facilitate high levels of gene expression in lungs of mice. However, despite significant improvement in gene transfer activity, we show here that the efficiency of GL-67-mediated gene transduction of intact airway epithelia is still relatively low. Administration of GL-67:pCF1-CFTR (encoding the cystic fibrosis transmembrane conductance regulator) complexes into the nasal epithelium of cystic fibrosis (CF) transgenic mice resulted only in marginal correction of the ion transport defects. Measurements of nasal potential differences (PD) showed no correction of the sodium (Na) transport defect, and only partial restitution of the chloride (Cl) transport defect was achieved in a small proportion of the animals after perfusion of the nasal epithelium with the complexes. Furthermore, in contrast to results obtained following instillation of GL-67:pDNA complexes into the lungs of mice, perfusion of GL-67:pDNA into the nasal epithelium resulted only in a moderate enhancement of gene transduction activity relative to that attained with naked pDNA alone. To determine the basis for this low efficiency of transfection, a series of studies was conducted to identify some of the barriers governing cationic lipid-mediated gene transfer to the airway epithelium. We show here that the transfection activity of GL-67 was affected by the polarization, differentiation, and proliferative state of the cells. Diminished transfection activity was observed with nonmitotic, highly polarized and differentiated airway epithelial cells. This observed reduction in gene expression with nonmitotic cells was determined to be due in part to inefficient nuclear translocation of the pDNA from the cytoplasm. Together these data indicate that much improvement in the ability of cationic lipids to transfect polarized and differentiated airway epithelial cells is a necessary prerequisite for effective cationic lipid-mediated gene therapy of airway diseases such as CF.

Published
1998
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49. Identification of an evolutionarily conserved 110 base-pair cis-acting regulatory sequence that governs Wnt-1 expression in the murine neural plate

Author

Rowitch, David H., Echelard, Yann, Danielian, Paul S., Gellner, Klaus, Brenner, Sydney, and McMahon, Andrew P.

Abstract

The generation of anterior-posterior polarity in the vertebrate brain requires the establishment of regional domains of gene expression at early somite stages. Wnt-1 encodes a signal that is expressed in the developing midbrain and is essential for midbrain and anterior hindbrain development. Previous work identified a 5.5 kilobase region located downstream of the Wnt-1 coding sequence which is necessary and sufficient for Wnt-1 expression in vivo. Using a transgenic mouse reporter assay, we have now identified a 110 base pair regulatory sequence within the 5.5 kilobase enhancer, which is sufficient for expression of a lacZ reporter in the approximate Wnt-1 pattern at neural plate stages. Multimers of this element driving Wnt-1 expression can partially rescue the midbrain-hindbrain phenotype of Wnt-1−/− embryos. The possibility that this region represents an evolutionarily conserved regulatory module is suggested by the identification of a highly homologous region located downstream of the wnt-1 gene in the pufferfish (Fugu rubripes). These sequences are capable of appropriate temporal and spatial activation of a reporter gene in the embryonic mouse midbrain; although, later aspects of the Wnt-1 expression pattern are absent. Genetic evidence has implicated Pax transcription factors in the regulation of Wnt-1. Although Pax-2 binds to the 110 base pair murine regulatory element in vitro, the location of the binding sites could not be precisely established and mutation of two putative low affinity sites did not abolish activation of a Wnt-1 reporter transgene in vivo. Thus, it is unlikely that Pax proteins regulate Wnt-1 by direct interactions with this cis-acting regulatory region. Our analysis of the 110 base pair minimal regulatory element suggests that Wnt-1 regulation is complex, involving different regulatory interactions for activation and the later maintenance of transgene expression in the dorsal midbrain and ventral diencephalon, and at the midbrain-hindbrain junction.

Published
1998
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