Quantitative description of the phase-separation behavior of the multivalent SLP65-CIN85 complex

Area of Science:

• Cell Biology
• Biochemistry
• Immunology

Background:

• Biomolecular condensates are crucial for cellular compartmentalization, complementing membrane-bound organelles.
• SLP65 and CIN85 are key effector proteins in B-cell antigen receptor (BCR) signaling, essential for effective immune responses.
• These proteins form phase-separated condensates with intracellular vesicles before antigen stimulation, priming B cells for activation.

Purpose of the Study:

• To investigate the molecular interactions between SLP65 and CIN85 proteins within phase-separated condensates.
• To determine the thermodynamic properties governing the formation and behavior of these condensates.
• To model and predict condensate formation using computational methods based on individual interaction strengths.

Main Methods:

• Quantified the dissociation constants of 18 individual interactions between proline-rich motifs (PRMs) of SLP65 and SH3 domains of CIN85.
• Measured the phase-separation properties of native and engineered SLP65/CIN85 systems.
• Utilized the LASSI (LAttice simulation engine for Sticker and Spacer Interactions) program to model condensate behavior.

Main Results:

• The thermodynamic properties of individual SLP65-CIN85 interactions were determined.
• Phase-separation properties of natural and designer SLP65/CIN85 constructs were accurately measured.
• LASSI modeling successfully reproduced experimental phase-separation behavior and identified a previously unknown intramolecular interaction.

Conclusions:

• Thermodynamic data of individual protein interactions can faithfully model the phase-separation behavior of the SLP65/CIN85 system.
• This study provides a quantitative framework for understanding condensate formation in immune signaling.
• The findings highlight the importance of multivalent interactions in cellular organization and function.