Phosphorylation of class II transactivator regulates its interaction ability and transactivation function

Area of Science:

• Immunology
• Molecular Biology
• Gene Regulation

Background:

• The MHC class II transactivator (CIITA) is essential for adaptive immune responses.
• CIITA orchestrates MHC class II gene expression by forming enhanceosome complexes with DNA-binding and co-activator proteins.

Purpose of the Study:

• To investigate the role of CIITA phosphorylation in regulating its interactions and transactivation potential.
• To identify the kinase responsible for CIITA phosphorylation and the specific sites involved.

Main Methods:

• In vitro phosphorylation assays using cAMP-dependent protein kinase (PKA).
• Analysis of CIITA interactions with co-activators (p300, RFX5) and self-association.
• Reporter gene assays to measure MHC class II promoter activity using wild-type and mutant CIITA.

Main Results:

• Hyper-phosphorylated CIITA interacts with p300, RFX5, and itself, enhancing MHC class II promoter activity.
• The C-terminal leucine-rich repeat (LRR) domain regulates CIITA self-association, negatively impacted by phosphorylation.
• PKA phosphorylates serine residues in CIITA, and this phosphorylation is critical for maximal transactivation, as shown by reduced activity in serine mutants.

Conclusions:

• CIITA phosphorylation status dictates its protein interactions and regulatory functions.
• PKA-mediated phosphorylation of specific serine residues is essential for CIITA's role in MHC class II gene expression and adaptive immunity.