Abstract
Extracellular matrix (ECM) plays a critical role in normal development, tissue repair, and is altered in several disease states. During these processes, expression of ECM genes is regulated temporally and spatially. Identification of regulatory elements that direct tissue- and stage-specific expression is important for understanding of the role of ECM genes in cellular differentiation and function. A significant number of ECM genes have been cloned and their regulatory elements have been identified. Reporter gene systems are widely used to study gene regulation and function (1 -3). In this chapter, we describe methods for the identification of promoters and enhancers of cartilage collagen genes for the α1(II) and α2(XI) chains using transgenic mice and transient transfection in cell cultures (4 -9).
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Gorman, C. M., Moffat, L. F., and Howard, B. H. (1982) Recombinant genomes which express chloramphenicol acetyltransferase in mammalian cells. Mol. Cell. Biol. 2, 1044–1051.
Neumann, J. R., Morency, C. A., and Russian, K. O. 1996 A novel assay for chloramphenicol acetyltransferase gene expresssion. BioTechniques 5, 444–447.
Norstedt. G., Enberg, B., Francis, S., Hansson, A., Hulthen, A., Lobie, P. E., et al. (1994) Cell transfection as a tool to study growth hormone action. Proc. Soc. Exp. Biol. Med. 206, 181–184.
Mukhopadhyay, K., Lefebvre, V., Zhou, G., Garofalo, S., Kimura, J. H., and de Crombrugghe, B. (1995) Use of a new rat chondrosarcoma cell line to delineate a 119-base pair chondrocyte-specific enhancer element and to define active promoter segments in the mouse pro-α1(II) collagen gene. J. Biol. Chem. 270, 17,711–17,719.
Krebsbach, P. H., Nakata, K., Bernier, S. M., Hatano, O., Miyashita, T., Rhodes, C. S., and Yamada, Y. (1996) Identification of a minimum enhancer sequence for the type II collagen gene reveals several core sequence motifs in common with the link protein gene. J. Biol. Chem. 271, 4298–4303.
Tsumaki, N., Kimura, T., Matsui, Y., Nakata, K., and Ochi, T. (1996) Separable cis-regulatory elements that contribute to tissue-and site-specific α2(XI) collagen gene expression in the embryonic mouse cartilage. J. Biol. Chem. 134, 1573–1582.
Bridgewater, L. C., Lefebvre, V., and de Crombrugghe, B. (1998) Chondrocytes-specific enahncer elements in the Col11α2 gene resemble the Col2a1 tissue-spe-cific enhancer. J. Biol. Chem. 273, 14,998–15,006.
Tsumaki, N., Kimura, T., Tanaka, K., Kimura, J. H., Ochi, H., and Yamada, Y. (1998) Modular arrangement of cartilage-and neural tissue-specific cis-elements in the mouse α2(XI) collagen promoter. J. Biol. Chem., in press.
Leung, K. K. H., Ng, L. J., Ho, K. Y., Tam, P. P. L., and Cheach K. S. E. (1998) Different cis-regulatory DNA elements mediate developmental stage-and tissue-specficif expression of the human COL2A1 gene in transgenic mice. J. Cell Biol. 141, 1291–1300.
Johnston, S. A. and Tang, D. C. (1997) Gene gun transfection of animal cells and genetic immunization. Meth. Cell. Biol. 43, 353–365.
Bernasconi, A. G., Rebuffat, A. G., Lovati, E., Frey, B. M., Frey, F. J., and Galli, I. (1997) Cortisol increases transfection efficiency of cells. FEBSLett. 419, 103–106.
Tseng, W. C., Haselton, F. R., and Giorgio, T. D. (1997) Transfection by cationic liposomes using simultaneous single cell measurements of plasmid delivery and transgene expression. J. Biol. Chem. 272, 25641–25647.
Neville, C., Rosenthal, N., and Hauschka, S. (1997) DNA transfection of cultured muscle cells. Meth. Cell. Biol. 52, 405–422.
Osborne, B. A., Smith, S. W., Liu, Z. G., McLaughlin, K. A., and Schwartz, L. M. (1995) Transient transfection assays to examine the requirement of putative cell death genes. Meth. Cell. Biol. 46, 99–106.
Hogan, B., Beddington, R., Constantini, F., and Lacy, E. (1994) Manipulating the Mouse Embryo: A Laboratory Manual, 2nd Ed. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY.
Bonnerot, C. and Nicolas, J.-F. (1993) Application of lacZ gene fusions to postimplantaion development, in Guide to Techniques in Mouse Development (Wassarman, P. M. and Depamphilis, M. L., ed.), Academic, San Diego, CA, pp. 451–469.
Hanley, T. and Merlie, J. P. (1991) Transgene detection in unpurified mouse tail DNA by polymerase chain reaction. BioTechniques 10, 56.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2000 Humana Press Inc.
About this protocol
Cite this protocol
Tsumaki, N., Liu, Y., Yamada, Y., Krebsbach, P. (2000). Enhancer Analysis of the α1(II) and α2(XI) Collagen Genes in Transfected Chondrocytes and Transgenic Mice. In: Streuli, C.H., Grant, M.E. (eds) Extracellular Matrix Protocols. Methods in Molecular Biology™, vol 139. Humana Press. https://doi.org/10.1385/1-59259-063-2:187
Download citation
DOI: https://doi.org/10.1385/1-59259-063-2:187
Publisher Name: Humana Press
Print ISBN: 978-0-89603-624-6
Online ISBN: 978-1-59259-063-6
eBook Packages: Springer Protocols