Future Prospects

We have shown that biotinylation tagging is highly efficient in cultured cells (Fig. 1) and transgenic mice (1), and we have used this approach to identify a number of different complexes formed by the essential hematopoietic transcription factor GATA-1 (6). Owing to its efficiency and ease of application, biotinylation tagging offers the prospect of rapidly expanding the characterization of transcription factor complexes. For example, the biotinylation tagging of the hematopoietic transcription factor partners of GATA-1 and the characterization of their protein complexes will lead to the rapid elucidation of the distinct and overlapping transcriptional networks these factors regulate in hematopoie-sis. Similarly, the biotinylation tagging of chromatin cofactors will lead to a better understanding of their interactions with tissue-specific transcription factors and the molecular basis of their functions (i.e., chromatin remodeling and modification in activation and repression). Furthermore, efforts in reducing the background along the lines described here (i.e., a prepurification steps such as gel filtration or the use of protease cleavage) will help in further expanding the utility of biotinylation tagging, for example, in preserving the native properties of complexes or in determining stoichiometries. The utility of biotinyla-tion tagging will be further increased through the development of additional tools such as the recent derivation of a transgenic mouse strain that expresses BirA ubiquitously in all tissues (8), or the construction of a codon-optimized version of BirA for the efficient expression in mammalian cells (9). The recent description of the biotinylation of cell surface proteins (10) should also serve to expand the utility of this approach. Lastly, it should be noted that in vivo biotinylation tagging can also be employed (e.g., instead of antibodies) in all other applications involving an affinity purification or detection step, such as immunofluorescence (1), immunoprecipitation (1,11), and chromatin immuno-precipitation (ChIP) assays (1,12).

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