This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Haas, D. L.
Right arrow Articles by Kohn, D. B.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Haas, D. L.
Right arrow Articles by Kohn, D. B.

 Previous Article  |  Next Article 

Journal of Virology, September 2003, p. 9439-9450, Vol. 77, No. 17
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.17.9439-9450.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

The Moloney Murine Leukemia Virus Repressor Binding Site Represses Expression in Murine and Human Hematopoietic Stem Cells

Dennis L. Haas, Carolyn Lutzko, Aaron C. Logan, Gerald J. Cho, Dianne Skelton, Xiao Jin Yu, Karen A. Pepper, and Donald B. Kohn*

Division of Research Immunology/BMT, Children’s Hospital Los Angeles, and the Departments of Pediatrics and Molecular Microbiology & Immunology, Keck School of Medicine, University of Southern California, Los Angeles, California

Received 11 March 2003/ Accepted 6 June 2003

The Moloney murine leukemia virus (MLV) repressor binding site (RBS) is a major determinant of restricted expression of MLV in undifferentiated mouse embryonic stem (ES) cells and mouse embryonal carcinoma (EC) lines. We show here that the RBS repressed expression when placed outside of its normal MLV genome context in a self-inactivating (SIN) lentiviral vector. In the lentiviral vector genome context, the RBS repressed expression of a modified MLV long terminal repeat (MNDU3) promoter, a simian virus 40 promoter, and three cellular promoters: ubiquitin C, mPGK, and hEF-1a. In addition to repressing expression in undifferentiated ES and EC cell lines, we show that the RBS substantially repressed expression in primary mouse embryonic fibroblasts, primary mouse bone marrow stromal cells, whole mouse bone marrow and its differentiated progeny after bone marrow transplant, and several mouse hematopoietic cell lines. Using an electrophoretic mobility shift assay, we show that binding factor A, the trans-acting factor proposed to convey repression by its interaction with the RBS, is present in the nuclear extracts of all mouse cells we analyzed where expression was repressed by the RBS. In addition, we show that the RBS partially repressed expression in the human hematopoietic cell line DU.528 and primary human CD34+ CD38- hematopoietic cells isolated from umbilical cord blood. These findings suggest that retroviral vectors carrying the RBS are subjected to high rates of repression in murine and human cells and that MLV vectors with primer binding site substitutions that remove the RBS may yield more-effective gene expression.

* Corresponding author. Mailing address: Division of Research Immunology/BMT, Children’s Hospital Los Angeles, 4650 Sunset Blvd., Los Angeles, CA 90027. Phone: (323) 669-4617. Fax: (323) 667-1021. E-mail: dkohn{at}chla.usc.edu.

Journal of Virology, September 2003, p. 9439-9450, Vol. 77, No. 17
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.17.9439-9450.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Wolf, D., Hug, K., Goff, S. P. (2008). TRIM28 mediates primer binding site-targeted silencing of Lys1,2 tRNA-utilizing retroviruses in embryonic cells. Proc. Natl. Acad. Sci. USA 105: 12521-12526 [Abstract] [Full Text]  
  • Hendrickson, B., Senadheera, D., Mishra, S., Bui, K. C. T., Wang, X., Chan, B., Petersen, D., Pepper, K., Lutzko, C. (2007). Development of Lentiviral Vectors with Regulated Respiratory Epithelial Expression In Vivo. Am. J. Respir. Cell Mol. Bio. 37: 414-423 [Abstract] [Full Text]  
  • Luo, P., Wang, A., Payne, K. J., Peng, H., Wang, J.-g., Parrish, Y. K., Rogerio, J. W., Triche, T. J., He, Q., Wu, L. (2007). Intrinsic Retinoic Acid Receptor {alpha}-Cyclin-Dependent Kinase-Activating Kinase Signaling Involves Coordination of the Restricted Proliferation and Granulocytic Differentiation of Human Hematopoietic Stem Cells. Stem Cells 25: 2628-2637 [Abstract] [Full Text]  
  • Camara-Clayette, V., Le Pesteur, F., Vainchenker, W., Sainteny, F. (2006). Quantitative Oct4 Overproduction in Mouse Embryonic Stem Cells Results in Prolonged Mesoderm Commitment During Hematopoietic Differentiation In Vitro. Stem Cells 24: 1937-1945 [Abstract] [Full Text]  
  • Lagresle-Peyrou, C., Yates, F., Malassis-Seris, M., Hue, C., Morillon, E., Garrigue, A., Liu, A., Hajdari, P., Stockholm, D., Danos, O., Lemercier, B., Gougeon, M.-L., Rieux-Laucat, F., de Villartay, J.-P., Fischer, A., Cavazzana-Calvo, M. (2006). Long-term immune reconstitution in RAG-1-deficient mice treated by retroviral gene therapy: a balance between efficiency and toxicity. Blood 107: 63-72 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»