Abstract
The mammalian epidermis is a self-renewing stratified squamous epithelium. Its basal cell layer contains proliferating keratinocytes that exit the cell cycle when they move into the suprabasal compartment. These cells activate a gene differentiation program aimed at building a protective epidermal barrier as they move toward the surface, successively going through the spinous and granular layers. At the completion of this process, the keratinocytes become enucleated and form the cornified layer, the surface layer of the skin. The highly cross-linked protein–lipid envelope and extracellular lipids in the cornified layer along with cell–cell adhesions in the granular layer are required for an effective epidermal barrier. Transcriptional mechanisms are critical for the formation of the epidermal barrier, and in this chapter, we describe methods to evaluate the role of a transcription factor (TF) in epidermal differentiation. To identify direct target genes of a TF, we propose a combination of bioinformatics and experimental approaches. The ultimate goal of these approaches is to understand the mechanisms whereby a TF regulates epidermal barrier formation.
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References
Nickoloff, B. J. (2006) Keratinocytes regain momentum as instigators of cutaneous inflammation. Trends Mol Med 12, 102–6.
Blanpain, C., and Fuchs, E. (2009) Epidermal homeostasis: a balancing act of stem cells in the skin. Nat Rev Mol Cell Biol 10, 207–17.
Koster, M. I., and Roop, D. R. (2007) Mechanisms regulating epithelial stratification. Annu Rev Cell Dev Biol 23, 93–113.
Long, A. D., Mangalam, H. J., Chan, B. Y., Tolleri, L., Hatfield, G. W., and Baldi, P. (2001) Improved statistical inference from DNA microarray data using analysis of variance and a Bayesian statistical framework. Analysis of global gene expression in Escherichia coli K12. J Biol Chem 276, 19937–44.
Yu, Z., Mannik, J., Soto, A., Lin, K. K., and Andersen, B. (2009) The epidermal differentiation-associated Grainyhead gene Get1/Grhl3 also regulates urothelial differentiation. Embo J 28, 1890–903.
Lin, K. K., Chudova, D., Hatfield, G. W., Smyth, P., and Andersen, B. (2004) Identification of hair cycle-associated genes from time-course gene expression profile data by using replicate variance. Proc Natl Acad Sci USA 101, 15955–60.
Lin, K. K., Kumar, V., Geyfman, M., Chudova, D., Ihler, A. T., Smyth, P., Paus, R., Takahashi, J. S., and Andersen, B. (2009) Circadian clock genes contribute to the regulation of hair follicle cycling. PLoS Genet 5, e1000573.
Saeed, A. I., Bhagabati, N. K., Braisted, J. C., Liang, W., Sharov, V., Howe, E. A., Li, J., Thiaga-rajan, M., White, J. A., and Quackenbush, J. (2006) TM4 microarray software suite. Methods Enzymol 411, 134–93.
Saeed, A. I., Sharov, V., White, J., Li, J., Liang, W., Bhagabati, N., Braisted, J., Klapa, M., Currier, T., Thiagarajan, M., Sturn, A., Snuffin, M., Rezantsev, A., Popov, D., Ryltsov, A., Kostukovich, E., Borisovsky, I., Liu, Z., Vinsavich, A., Trush, V., and Quackenbush, J. (2003) TM4: a free, open-source system for microarray data management and analysis. Biotechniques 34, 374–8.
Sandelin, A., Wasserman, W. W., and Lenhard, B. (2004) ConSite: web-based prediction of regulatory elements using cross-species comparison. Nucleic Acids Res 32, W249–52.
Vega, V. B., Bangarusamy, D. K., Miller, L. D., Liu, E. T., and Lin, C. Y. (2004) BEARR: Batch Extraction and Analysis of cis-Regulatory Regions. Nucleic Acids Res 32, W257–60.
Chi, X., Zhang, S. X., Yu, W., DeMayo, F. J., Rosenberg, S. M., and Schwartz, R. J. (2003) Expression of Nkx2-5-GFP bacterial artificial chromosome transgenic mice closely resembles endogenous Nkx2-5 gene activity. Genesis 35, 220–6.
Decker, T., Pasca di Magliano, M., McManus, S., Sun, Q., Bonifer, C., Tagoh, H., and Busslinger, M. (2009) Stepwise activation of enhancer and promoter regions of the B cell commitment gene Pax5 in early lymphopoiesis. Immunity 30, 508–20.
Wells, J., and Dai, X. Using siRNA knockdown in HaCaT cells to study transcriptional control of epidermal proliferation potential. Methods Mol Biol 585, 107–25.
Choe, S. E., Boutros, M., Michelson, A. M., Church, G. M., and Halfon, M. S. (2005) Preferred analysis methods for Affymetrix GeneChips revealed by a wholly defined control dataset. Genome Biol 6, R16.
Wang, N., Lin, K. K., Lu, Z., Lam, K. S., Newton, R., Xu, X., Yu, Z., Gill, G. N., and Andersen, B. (2007) The LIM-only factor LMO4 regulates expression of the BMP7 gene through an HDAC2-dependent mechanism, and controls cell proliferation and apoptosis of mammary epithelial cells. Oncogene 26, 6431–41.
Yu, Z., Lin, K. K., Bhandari, A., Spencer, J. A., Xu, X., Wang, N., Lu, Z., Gill, G. N., Roop, D. R., Wertz, P., and Andersen, B. (2006) The Grainyhead-like epithelial transactivator Get-1/Grhl3 regulates epidermal terminal differentiation and interacts functionally with LMO4. Dev Biol 299, 122–36.
Acknowledgments
This work was supported by TRDRP dissertation award 17DT-0192 (to A.B.), NIH-NLM Biomedical Informatics Training grant 5T15LM007743 (to MS) and NIH grant AR44882 (to BA). The authors acknowledge the contributions of other laboratory members to the approaches described in this chapter. We thank Amy Soto and Suman Verma for reading the manuscript. We acknowledge the Expression Analysis Core at UC Davis for ChIP training.
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Bhandari, A., Salmans, M.L., Gordon, W., Andersen, B. (2011). Transcriptional Regulation of Epidermal Barrier Formation. In: Turksen, K. (eds) Permeability Barrier. Methods in Molecular Biology, vol 763. Humana Press. https://doi.org/10.1007/978-1-61779-191-8_3
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DOI: https://doi.org/10.1007/978-1-61779-191-8_3
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