Synthesis and assembly of spectrin during avian erythropoiesis: stoichiometric assembly but unequal synthesis of alpha and beta spectrin.
The synthesis and assembly of spectrin was investigated in erythroid cells during chicken embryo development. Immunoprecipitation of Triton X-100-soluble and -insoluble cytoskeletal fractions with alpha- and beta-spectrin antisera show that, at steady state, alpha and beta spectrin are present in stoichiometric amounts, and exclusively, in the cytoskeleton. However, pulse labeling of cells and in vitro translation of total erythroid cell RNA reveal that alpha spectrin is synthesized in a two to three fold excess over beta spectrin. Pulse-chase experiments show that newly synthesized alpha and beta spectrin are present in both the cytoskeletal and soluble fractions, and that stoichiometric amounts are stably assembled in the cytoskeleton. On the other hand, there is a severalfold excess of alpha relative to beta spectrin in the soluble fraction, both of which turn over with a half-life of 50 min. In cells from 4 day old embryos, more than 80% of the newly synthesized beta spectrin, but only 10% of the alpha spectrin, are present in the cytoskeleton. Thus, early in development, the association of alpha and beta spectrin with the membrane-cytoskeleton may be rate-limited by the amount of beta spectrin synthesized. Later on in erythroid development, progressively lesser proportions of newly synthesized beta spectrin are present in the cytoskeleton, suggesting that during development, the rate of association of beta spectrin with the membrane-cytoskeleton becomes limited by some other membrane-cytoskeletal component.
D000818:Animals; D017487:Ankyrins; D002458:Cell Fractionation; D002642:Chick Embryo; D003593:Cytoplasm / Q000032:analysis; D003599:Cytoskeleton / Q000378:metabolism*; D004912:Erythrocytes / Q000378:metabolism*; D004920:Erythropoiesis; D006461:Hemolysis; D008565:Membrane Proteins / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D013049:Spectrin / Q000096:biosynthesis / Q000378:metabolism*
D017487:Ankyrins; D008565:Membrane Proteins; D012333:RNA, Messenger; D013049:Spectrin