A single historical substitution drives an increase in acetylcholine receptor complexity

Area of Science:

• Biochemistry
• Molecular Biology
• Neuroscience

Background:

• Human adult muscle-type acetylcholine receptors (AChRs) are vital heteropentameric ion channels.
• Their complex subunit composition and stoichiometry are crucial for function but their evolutionary origins remain unclear.

Purpose of the Study:

• To investigate how evolutionary changes in subunits influence the stoichiometry of functional acetylcholine receptors.
• To understand the molecular mechanisms driving subunit entrenchment in heteromeric proteins.

Main Methods:

• Ancestral sequence reconstruction to recreate ancestral subunits.
• Single-channel electrophysiology to assess receptor function.
• Concatenated subunit expression to control assembly.

Main Results:

• An ancestral β-subunit could replace both extant β and neighboring δ-subunits in functional AChRs.
• A single amino acid substitution in the ancestral β-subunit re-established the requirement for a δ-subunit.
• This mutation drove an increase in receptor complexity and subunit entrenchment.

Conclusions:

• A single historical amino acid substitution can necessitate increased complexity in acetylcholine receptor assembly.
• Stepwise mutations can drive the entrenchment of subunit requirements in heteromeric proteins.
• This provides a model for understanding the evolution of protein complexity.