Setdb1 and Atf7IP form a hetero-trimeric complex that blocks Setdb1 nuclear export

Area of Science:

• Epigenetics
• Molecular Biology
• Structural Biology

Background:

• Histone H3K9 methylation (H3K9me) by Setdb1 silences retrotransposons (rTEs).
• Atf7IP is crucial for Setdb1's nuclear localization, activation, and chromatin recruitment.
• Structural details of the Setdb1/Atf7IP interaction remain largely uncharacterized.

Purpose of the Study:

• To elucidate the structural basis of the Setdb1/Atf7IP interaction.
• To understand how Atf7IP regulates Setdb1's nuclear localization and activity.
• To investigate the role of Atf7IP paralogs in Setdb1 complex formation and function.

Main Methods:

• AlphaFold2 predictions for structural modeling.
• Biochemical reconstitutions to validate complex formation.
• In vitro and in cell assays to study protein interactions.

Main Results:

• Setdb1 and Atf7IP form a stable heterotrimeric complex (1:2 stoichiometry) in vitro and in cells.
• Atf7IP self-associates, and Setdb1 binding resolves these multimeric complexes.
• Atf7IP binds Setdb1 via coiled-coil interactions, including Setdb1's nuclear export signals (NES), thereby preventing CRM1-mediated nuclear export.
• Setdb1 also forms heterotrimers with Atf7IP2, and mixed heterotrimers (Setdb1/Atf7IP/Atf7IP2) are possible.

Conclusions:

• Atf7IP directly binds Setdb1's NES motifs, inhibiting nuclear export and stabilizing Setdb1 in the nucleus.
• The formation of heterotrimers with varying Atf7IP/Atf7IP2 ratios allows for fine-tuning of Setdb1 activity and H3K9me.
• These findings provide structural insights into the regulation of heterochromatin formation by the Setdb1/Atf7IP complex.