HP1γ self-assembles and cooperates with KAP1 in repression of long noncoding RNA AI662270 in ESCs

Area of Science:

• Molecular biology
• Epigenetics
• Structural biology

Background:

• Heterochromatin protein 1 (HP1) family members regulate gene expression and heterochromatin assembly.
• The chromoshadow domain (CSD) of HP1 proteins mediates interactions with various partners.
• Understanding HP1 interactions is key to deciphering transcriptional regulation.

Purpose of the Study:

• To elucidate the molecular mechanisms of HP1γ chromoshadow domain (HP1γ_CSD) binding to KAP1.
• To investigate HP1γ self-assembly and compare it with HP1α.
• To determine the in vivo functional significance of HP1γ-KAP1 interactions in transcriptional repression.

Main Methods:

• X-ray crystallography and Nuclear Magnetic Resonance (NMR) spectroscopy to determine complex structures.
• Mass photometry to analyze HP1γ_CSD self-assembly.
• In vivo rescue experiments in KAP1- and HP1-knockout mouse embryonic stem cells.

Main Results:

• HP1γ_CSD forms a stable dimer of dimers via antiparallel assembly, recognizing the KAP1 HP1 box (KAP1_Hbox).
• HP1α_CSD exhibits concentration-dependent parallel oligomerization, distinct from HP1γ_CSD.
• Electrostatic interactions stabilize the HP1γ_CSD dimer interface, contrasting with HP1α_CSD's hydrophobic interface.
• HP1γ and KAP1 cooperatively repress the long noncoding RNA AI662270 in vivo.

Conclusions:

• HP1γ_CSD and HP1α_CSD display distinct mechanisms of self-assembly and interaction with KAP1.
• The unique HP1γ-KAP1 interaction is essential for the specific repression of lncRNA AI662270.
• Cellular functions of HP1 proteins are not redundant, with HP1γ playing a specialized role.