Modular organization of the E2F1 activation domain and its interaction with general transcription factors TBP and TFIIH

Abstract

The transcriptional activator E2F1 regulates the expression of genes at the G1/S boundary. We have characterized interactions of the E2F1 activation domain with two general transcription factors, the TATA-box binding protein (TBP) and TFIIH. Two distinct binding sites on E2F1 were identi®ed for TBP (amino acids 386 ± 417 and 415 ± 437) each of which supported activation in mammalian cells when expressed as a fusion to a heterologous DNA-binding domain. Neither of these minimal activation domains independently bound TFIIH; rather, the TFIIH binding site of E2F1 overlaps both domains. Loss of TFIIH-binding by E2F1 resulted in a 60 ± 65% reduction in transactivation, suggesting that the E2F1/TFIIH interaction is important, but not essential, for transactivation. The retinoblastoma protein (Rb) binds directly to E2F1 and represses E2F1-mediated transactivation. We have demonstrated that recombinant Rb can compete with TBP and the p62 subunit of TFIIH for binding to immobilized E2F1. A tumorigenic form of Rb de®cient in repressing E2F1-mediated transactivation is likewise de®cient in displacing TBP from E2F1. We propose that competition between Rb and both TBP and TFIIH for binding to E2F1 is a mechanism by which Rb inhibits transactivation by E2F1.

Structural basis for the recognition of the E2F transactivation domain by the retinoblastoma tumor suppressor

“…The molecular mechanism for E2F-driven transcription activation is unclear at present. However, the transactivation domain of E2F-1 is known to interact with general transcription factors such as TATA-boxbinding protein (TBP;Hagemeier et al 1993;Pearson and Greenblatt 1997), TFIIA/TFIIB (Ross et al 1999), and the coactivator p300/CBP Fry et al 1999).…”

Functional analysis of a promoter variant identified in the CFTR gene in cis of a frameshift mutation

“…Indeed, E2F and Sp1 have been shown to interact with several proteins of the general transcription machinery, such as TFIIH and TFIID. [22][23][24][25] These data suggest that the binding of these factors putatively modifies the use of the major TSS essential for CFTR transcription during lung development. 26,27 The identification of alterations in non-coding regions and the determination of the impact of each allele component have several important consequences for recessive disorders such as CF.…”

E2F4 (E2F transcription factor 4, p107/p130-binding)

“…E2F4 interacts primarily with p130/RBL2, p107/RBL1 and to a lesser extent with Rb/RB1 (Beijersbergen et al, 1994;Ginsberg et al, 1994;Ikeda et al, 1996;Moberg et al, 1996;Li et al, 1997). Pocket proteins modulate E2F transcription factor activity via two different mechanisms: 1-by preventing general transcription machinery and chromatin-remodeling protein recruitment (Helin et al, 1992;Flemington et al, 1993;Hagemeier et al, 1993;Helin et al, 1993a;Pearson and Greenblatt., 1997) and 2-by actively repressing gene transcription (Harbour and Dean, 2000;Singh et al, 2010). In fact, pocket proteins have been shown to recruit histone deacetylase enzymes (HDACs) (Brehm et al, 1998;Luo et al, 1998;Dahiya et al, 2000), the histone methyltransferase SUV39H1 (Nielsen et al, 2001;Vandel et al, 2001), SWI/SNF family members (BRG1, Brm) (Dunaief et al, 1994;Singh et al, 1995;Strobeck et al, 2000;Zhang et al, 2000;Iakova et al, 2003), the Sin3B repressor complex (via RBP1 and SAP30) Grandinetti and David., 2008) and the ErbB3 binding protein Ebp1 (Zhang et al, 2003), all of which contribute to chromatin compaction and thus, to transcriptional repression (Kouzarides, 2007).…”

“…E2F transcription factors can bind DNA as homodimers although to a much lesser extent than heterodimers (Huber et al, 1993). Transactivation Domain: The transactivation domain mediates interaction with transcriptional machinery, co-activators and chromatinremodeling proteins (Emili and Ingles, 1995;Trouche and Kouzarides, 1996;Pearson and Greenblatt, 1997;McMahon et al, 1998;Ross et al, 1999;Vandel and Kouzarides, 1999;Martinez-Balbas et al, 2000;Marzio et al, 2000;Lang et al, 2001;Louie et al, 2004;Taubert et al, 2004) and includes the Pocket Protein Binding Domain. E2F4 interacts with p130/RBL2, p107/RBL1 and to a lesser extent with pRb/RB1 (Beijersbergen et al, 1994;Ginsberg et al, 1994;Ikeda et al, 1996;Moberg et al, 1996;Li et al, 1997).…”

“…Binding specificity of E2F transcription factors to different target gene promoters is affected by the DNA sequence itself, E2F transcription factors, DP partners as well as other factors (Karlseder et al, 1996;Lin et al, 1996;Shin et al, 1996;Wells et al, 1997;Le Cam et al, 1999;Chen et al, 2002;Schlisio et al, 2002;Araki et al, 2003;Giangrande et al, 2003;Giangrande et al, 2004;Zhu et al, 2004). The transactivation domain of E2F transcription factors, including E2F4, mediates target gene transcription through two distinct mechanisms: 1-by recruiting general transcription machinery such as TBP/TFIID, TFIIA and TFIIH which promote RNA pol II pre-initiation complex (PIC) assembly (Hagemeier et al, 1993;Emili and Ingles, 1995;Pearson and Greenblatt, 1997;Ross et al, 1999;Vandel and Kouzarides., 1999) and 2-by relaxing chromatin structure at promoters by interacting with histone acetyltransferases (HAT) such as Tip60 (Taubert et al, 2004), p300/CBP (Trouche and Kouzarides, 1996;Trouche et al, 1996;Martinez-Balbas et al, 2000;Marzio et al, 2000) and PCAF/GCN5 (MartinezBalbas et al, 2000;Marzio et al, 2000;Lang et al, 2001). The transactivation domain also includes a pocket protein interacting domain.…”