Endoplasmic Reticulum Stress-responsive Transcription Factor ATF6α Directs Recruitment of the Mediator of RNA Polymerase II Transcription and Multiple Histone Acetyltransferase Complexes

Abstract

Background: Transcription factor ATF6␣ is a master regulator of genes induced by endoplasmic reticulum stress. Results: ATF6␣ can recruit Mediator and histone acetyltransferase complexes to promoter DNA via interactions with overlapping sites in the activation domain of ATF6␣. Conclusion: ATF6␣ sequences essential for gene activation recruit Mediator and histone acetyltransferases. Significance: Learning how coregulators communicate with DNA binding transcription factors is important for understanding gene regulation.

Role for Human Mediator Subunit MED25 in Recruitment of Mediator to Promoters by Endoplasmic Reticulum Stress-responsive Transcription Factor ATF6α

“…The ATF6␣ AD includes an eight amino acid sequence that is required for optimal ATF6␣-dependent transcription activation in cells (30) and for binding to the Mediator complex in vitro (12). This sequence resembles a VP16 AD region referred to as VN8, which is essential for VP16 transcription activity and Mediator binding (13,31,32).…”

“…Expression and Purification of Recombinant Proteins-GST-ATF6␣-AD (ATF6␣ amino acids 1-150) was expressed in Escherichia coli and purified as described (12). The MED25-VBD (MED25 amino acids 394 -543) was cloned into pET21a with a C-terminal 6-histidine tag and transformed into BL-21 DE3 cells.…”

“…The MED25-VBD (MED25 amino acids 394 -543) was cloned into pET21a with a C-terminal 6-histidine tag and transformed into BL-21 DE3 cells. Protein expression and purification were conducted as described (12) with the following modifications. Cells were harvested at 6000 ϫ g and resuspended in 20 ml of 50 mM Tris-Cl, pH 7.9, 300 mM NaCl, 10% glycerol, 1:1000 protease inhibitor mixture (Sigma), and 20 mM ␤-mercaptoethanol.…”

“…Immobilized DNA binding assays were performed essentially as described (12). Briefly, saturating amounts of GAL4-DNA binding domain or GAL4 fusion proteins were incubated in binding buffer containing 20 mM Hepes-NaOH pH 7.9, 80 mM NaCl, 10% glycerol, 0.2 mM EDTA, 0.02% Nonidet P-40, 0.1 mg/ml BSA, and 1 mM DTT with 400 fmol of a biotinylated DNA fragment containing 5 GAL4 DNA binding sites upstream of the adenovirus major late promoter and immobilized on Dynabeads M-280 (Invitrogen) streptavidin magnetic beads.…”

Endoplasmic reticulum proteostasis in glioblastoma—From molecular mechanisms to therapeutic perspectives

“…In contrast to HDACs, the Spt-Ada-Gcn5-acetyltransferase (SAGA) HAT complex is essential for ER stress-induced gene transcription (85). For instance, ATF6 activates transcription of ER stress response genes through the recruitment of several HAT complexes including SAGA and Ada2a-containing (ATAC) molecules (86,87). Transcription of ATF4 target genes CHOP, ATF3, and JMJD3 requires HAT activity (79).…”