Erk signaling and chromatin remodeling in MMTV promoter activation by progestins

Abstract

Transcription from the mouse mammary tumor virus (MMTV) promoter can be induced by progestins. The progesterone receptor (PR) binds to a cluster of five hormone responsive elements (HREs) and activates the promoter by synergistic interactions with the ubiquitous transcription factor, nuclear factor 1 (NF1). Progesterone treatment of cells in culture leads to activation of the Src/Ras/Erk/Msk1 cascade. Selective inhibition of Erk, or its target kinase Msk1, interferes with chromatin remodeling and blocks MMTV activation. A complex of activated PR, Erk and Msk1 is recruited to promoter after 5 min of hormone treatment and phosphorylates histone H3 at serine 10. This modification promotes the displacement of HP1gamma and subsequent chromatin remodeling. Progestin treatment leads to the recruitment of the BAF complex, which selectively displaces histones H2A and H2B from the nucleosome containing the HREs. The acetyltransferase PCAF is also required for induction of progesterone target genes and acetylates histone H3 at K14, an epigenetic mark, which interacts with Brg1 and Brm, anchoring the BAF complex to chromatin. In nucleosomes assembled on either MMTV or mouse rDNA promoter sequences, SWI/SNF displaces histones H2A and H2B from MMTV, but not from the rDNA nucleosome. Thus, the outcome of nucleosome remodeling by purified SWI/SNF depends on DNA sequence. The resultant H3/H4 tetramer particle is then the substrate for subsequent events in induction. Thus, initial activation of the MMTV promoter requires activation of several kinases and PCAF leading to phosphoacetylation of H3, and recruitment of BAF with subsequent removal of H2A/H2B.

Figure 1. Schematic representation of the main cis elements in the MMTV promoter and their occupancy in nucleosomes assembled in vitro (upper panel) and in intact cells after hormone induction (lower panel).

The positions covered by the main population of histone octamers are indicated by the grey ovals. The HREs, the NF1 binding site and the TATA box are indicated. The numbers refer to the distance in nucleotides from the transcription start site. The hormone receptor (PR) dimers are depicted in yellow and the NF1 dimer by green circles.

Figure 2. Hypothetical model for the initial steps of MMTV promoter induction.

Progestins bind to cytoplasmic PR/ER complexes, and activate the Src/Ras/Erk pathway, leading to nuclear accumulation of activated Erk. Most of PR is nuclear and complexed to chaperones (Hsps). Upon binding of progestins, PR homodimers dissociate from chaperones and a fraction of PR is phosphorylated by pErk, which also phosphorylates Msk1. A “PR -activated complex” composed of pPR/pErk/pMsk1 is formed.

Figure 3. Hypothetical model for the role of “PR activated complex” in chromatin.

Upper Panel: (1) An HP1γ-containing complex is bound to the promoter in the absence of hormone; (2) PR-activated complex binds BAF, but not PBAF; (3) PR-activated complex is recruited to the nucleosome B upon hormone induction, followed by H3 phosphoacetylation. The repressive complex is displaced from the promoter; (4) Recruitment of the BAF complex to the nucleosome B is stabilized by PCAF-dependent H3K14 acetylation, enabling ATP-dependent H2A/H2B displacement (and/or histone variants incorporation); (5) This metastable opening facilitates NF1 binding, generating a stable platform for the assembly of further PR molecules, BAF, coactivators and the basal transcriptional machinery, including RNA polymerase II. Lower Panel: T47D-MTVL cells were untreated (0) or treated for 5 min with 10 nM R-5020 and subjected to ChIP assays with the indicated specific antibodies. A schematic representation of the ChIPs described in (Vicent et al., 2006a) and (Vicent et al., 2009) is shown.