Abstract
Gene expression regulation by miRNAs has been reported to control key aspects of B cell differentiation and function (Chen et al., Science 303:83–86, 2004; Xiao et al., Cell 131:146–159, 2007; O’Carroll et al., Genes Dev. 21:1999–2004, 2007; Koralov et al. Cell 132:860–874, 2008; Rodriguez et al., Science 316:608–611, 2007; Costinean et al., Proc Natl Acad Sci USA 103:7024–7029, 2006; Thai et al., Science 316:604–608, 2007; Vigorito et al., Immunity 27:847–859, 2007; Dorsett et al., Immunity 28:630–638, 2008; Teng et al., Immunity 28:621–629, 2008; de Yebenes et al., J Exp Med 205:2199–2206, 2008; He et al., Nature 435:828–833, 2005; Ventura et al. Cell 132:875–886, 2008; Xiao et al., Nat Immunol 9:405–414, 2008). In this chapter, we describe the methodology used to perform a functional screening of a miRNA library to identify miRNAs relevant for mature B cell function in primary mouse B cells. These procedures include the construction of a miRNA library and the expression of individual miRNA clones in spleen B cells, as well as the description of functional assays used to determine the impact of miRNA expression on several aspects of B cell function, such as proliferation, apoptosis, and class switch recombination.
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Acknowledgments
We would like to thank Dr He for kindly sharing a miRNA library with us and for helpful discussion. This work was supported by grants from the Ministerio de Educación y Ciencia (SAF2007-63130), the Comunidad Autónoma de Madrid (DIFHEMAT-CM), and the European Research Council (BCLYM-207844). V. G. de Yebenes and A. R. Ramiro are supported by the Ramón y Cajal program from the Ministerio de Educación y Ciencia.
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de Yébenes, V.G., Ramiro, A.R. (2010). MicroRNA Activity in B Lymphocytes. In: Monticelli, S. (eds) MicroRNAs and the Immune System. Methods in Molecular Biology, vol 667. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-60761-811-9_12
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DOI: https://doi.org/10.1007/978-1-60761-811-9_12
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