Sphingomyelinase restricts the lateral diffusion of CD4 and inhibits human immunodeficiency virus fusion

Area of Science:

• Virology
• Cell Biology
• Biochemistry

Background:

• Sphingomyelinase treatment was previously shown to inhibit human immunodeficiency virus type 1 (HIV-1) entry.
• The precise mechanism by which sphingomyelinase inhibits HIV-1 entry remained unclear.

Purpose of the Study:

• To elucidate the mechanism by which sphingomyelinase inhibits HIV-1 entry.
• To investigate the effect of sphingomyelinase on the lateral mobility and interactions of HIV-1 receptors (CD4 and CCR5).

Main Methods:

• Measuring fluorescence recovery after photobleaching to assess lateral diffusion of CD4 and CCR5.
• Assessing gp120 binding, HIV-1 attachment, and endocytosis.
• Determining membrane disposition of HIV receptor proteins using Triton X-100 extraction.
• Antibody cross-linking of CD4 to restrict its diffusion.

Main Results:

• Sphingomyelinase treatment decreased CD4 lateral diffusion by 4-fold, while CCR5 diffusion remained unchanged.
• Sphingomyelinase did not affect gp120 binding, HIV-1 attachment, endocytosis, or receptor membrane disposition.
• Restriction of CD4 diffusion by antibody cross-linking also inhibited HIV infection.
• Sphingomyelinase inhibited HIV fusion after CD4 engagement, with maximal inhibition at short coculture times and low CD4 expression.

Conclusions:

• Sphingomyelinase inhibits HIV-1 entry by inducing CD4 clustering, which impedes coreceptor engagement.
• This CD4 clustering prevents subsequent fusion events necessary for viral entry.
• The findings suggest a novel therapeutic strategy targeting CD4-receptor interactions to inhibit HIV-1 infection.