Reorientation of membrane polypeptides during erythrocyte maturation.

Messenger RNA extracted from the erythroleukemic cell line K562 was translated in vitro and the translation products reacted with an antiserum raised against human erythrocyte ghosts. Polypeptides immunoprecipitated by the antiserum were characterized by SDS-polyacrylamide gel electrophoresis and fluorography. The antiserum immunoprecipitated polypeptides with nominal molecular weights of 37,000 (p37), 20,000 (p20), 19,000 (p19), 18,000 (p18), 14,000 (p14), 13,000 (p13), and 11,000 (p11) daltons. Since the antiserum was raised against antigenic determinants present on both the inner and outer surface of the red cell membrane, differential absorption of antiserum with intact red cells, or ghosts, was used to localize the translation products to the inner or outer membrane surface. Absorption was also used to determine if any of the immunoprecipitated translation products represented membrane markers for the erythroid lineage. Absorption of the antiserum with red cell ghosts removed all antibodies reacting with in vitro translation products. Absorption with intact cells from various lineages removed anti-p20 antibodies and did not absorb anti-p19 or anti-p18 antibodies. Absorption with intact cells from all lineages except mature erythrocytes absorbed anti-p37, anti-p14, and anti-p13 antibodies, suggesting that these antigens are expressed on the outer membrane surface. Mature erythrocytes were incapable of absorbing these antibody populations, suggesting a lineage-specific reorientation of these antigens in the membrane during erythropoiesis.