Transcription in eukaryotes notes
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Many eukaryotes are estimated to have 20,000–25,000 genes (see Table 16.4). Some of these are expressed (transcribed) in all cells all of the time, while others are expressed as cells enter a particular pathway of differentiation or as conditions in and around the cells change. In the early 1980s, transcription researchers primarily explored DNA–protein interactions in vitro. Research focused on the purification of sequencespecific DNA-binding proteins by affinity chromatography, analysis of the transcriptional activity of promoters by reporter gene assays, in vitro transcription assays that allowed the fractionation of the general transcription machinery, and assays such as electrophoretic gel mobility shift assays (EMSA) and DNase I footprinting for analysis of cis-acting DNA elements with trans-acting factors (see Chapter 9 for methods). By the late 1980s, many sequence-specific DNA-binding proteins had been identified, purified, and their genes cloned. Upon further study, it became clear that in addition to DNA–protein interactions, protein–protein interactions were of critical importance for regulating gene transcription.
This insight was followed closely by the realization in the early 1990s that chromatin structure, nuclear architecture, and cellular compartmentalization must also be taken into account. Sections within this chapter will cover
protein-coding gene regulatory elements, transcription factors and their DNA-binding motifs, the general transcription machinery and the mechanism of RNA polymerase II transcription, transcriptional coactivators and corepressors, including chromatin modification and remodeling complexes, and signal-mediated nuclear import and export of proteins involved in regulating gene transcription.
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