Alphavirus 6K proteins form ion channels

Area of Science:

• Virology
• Molecular Biology
• Biophysics

Background:

• Ross River virus and Barmah Forest virus are Australian arboviruses belonging to the Alphavirus genus.
• Alphavirus infections are characterized by increased cell permeability to monovalent cations and virion budding.
• Virally encoded ion channels are hypothesized to play a role in these infection mechanisms.

Purpose of the Study:

• To investigate the ion channel-forming capabilities of the 6K proteins from Ross River virus and Barmah Forest virus.
• To determine the functional properties and orientation of these viral ion channels.

Main Methods:

• Bacterial expression and purification of 6K proteins from Ross River virus and Barmah Forest virus.
• Incorporation of purified 6K proteins into planar lipid bilayers with defined orientation.
• Electrophysiological recordings to measure channel activity and conductances.
• Inhibition studies using antibodies targeting the N and C termini of the 6K protein.

Main Results:

• The 6K proteins of Ross River virus and Barmah Forest virus were demonstrated to form cation-selective ion channels in planar lipid bilayers.
• Defined orientation (N-terminal cis, C-terminal trans) of the inserted 6K proteins was achieved.
• Channel activity was reversibly inhibited by antibodies, confirming protein identity and function.
• Measured channel conductances ranged from 40-800 picosiemens, indicating potential for various oligomerization states.

Conclusions:

• The 6K proteins of Ross River virus and Barmah Forest virus function as cation-selective ion channels.
• These viral ion channels likely contribute to the characteristic cellular changes observed during alphavirus infections.
• The ability of 6K proteins to form channels with varying conductances suggests flexibility in their oligomeric assembly.