Calcium Contributes to Polarized Targeting of HIV Assembly Machinery by Regulating Complex Stability

Polarized or precision targeting of protein complexes to their destinations is fundamental to cellular homeostasis, but the mechanism underpinning directional protein delivery is poorly understood. Here, we use the uropod targeting HIV synapse as a model system to show that the viral assembly machinery Gag is copolarized with the intracellular calcium (Ca²⁺) gradient and binds specifically with Ca²⁺. Conserved glutamic/aspartic acids flanking endosomal sorting complexes required for transport binding motifs are major Ca²⁺ binding sites. Deletion or mutation of these Ca²⁺ binding residues resulted in altered protein trafficking phenotypes, including (i) changes in the Ca²⁺-Gag distribution relationship during uropod targeting and/or (ii) defects in homo/hetero-oligomerization with Gag. Mutation of Ca²⁺ binding amino acids is associated with enhanced ubiquitination and a decline in virion release via uropod protein complex delivery. Our data that show Ca²⁺-protein binding, via the intracellular Ca²⁺ gradient, represents a mechanism that regulates intracellular protein trafficking.