DYNAMIC INTERACTIONS OF THE ASIALOGLYCOPROTEIN RECEPTOR SUBUNITS WITHCOATED PITS - ENHANCED INTERACTIONS OF H2 FOLLOWING ASSOCIATION WITH H1

Lateral mobility studies comparing native and mutated membrane protein s, combined with treatments that alter clatbrin lattice structure, can measure membrane protein-coated pit interactions in intact cells (Fir e, E., Zwart, D., Roth, M.G., an Henis, Y.I. (1991) J. Cell Biol. 115, 1585-1594). We applied this approach to study the interactions of the H1 and H2 human asialoglycoprotein receptor subunits with coated pits . The lateral mobilities of singly expressed and coexpressed H1 and H2 B (the H2 species that reaches the cell surface) were measured by fluo rescence photobleaching recovery. They were compared with mutant prote ins, H1(5A) (Tyr-5 replaced by Ala) and H2(5A) (Phe-5 replaced by Ala) . While the mobile fraction of H1, H2B, and their mutants were similar , the lateral diffusion rate (measured by D, the lateral diffusion coe fficient) was significantly slower for H1, whether expressed alone or with H2B. Coexpression with H1 reduced D of H2B to that of H1. Disrupt ion of the clathrin lattices by hypertonic medium elevated D of H1, H1 (5A), H2B, and H2(5A) to the same final level, without affecting their mobile fractions. Cytosol acidification, which retains altered clathr in lattices attached to the membrane and prevents coated vesicle forma tion, immobilized part of the H1 molecules, reflecting stable entrapme nt in ''frozen'' coated pits. H1(5A), H2B, and H2(5A) were not affecte d: however, coexpression of H2B with H1 conferred the sensitivity to c ytosol acidification on H2B. Our results suggest that H1 lateral mobil ity is inhibited by dynamic interactions with coated pits in which Tyr -5 is involved. H2B resembles H1(5A) rather than H1, and its interacti ons with coated pits are weaker; efficient interaction of H2B with coa ted pits depends on complex formation with H1.