Abstract
Two tubulin variants, isolated from chicken brain and erythrocytes and known to have different peptide maps and electrophoretic properties, are demonstrated to exhibit different assembly properties in vitro: 1) erythrocyte tubulin assembles with greater efficiency (lower critical concentration, greater elongation rate) but exhibits a lower nucleation rate than brain tubulin, and 2) erythrocyte tubulin readily forms oligomers whose presence significantly retards the rate of elongation, suggesting that tubulin oligomers may also be important for determining the rate of assembly and the length of microtubules in erythrocytes. Erythrocyte tubulin isolated by cycles of in vitro assembly-disassembly is also demonstrated to contain a 67-kDa tau factor that greatly enhances microtubule nucleation but has little effect on elongation rates or critical concentration. Immunofluorescence microscopy with tau antibody indicates that tau is specifically associated with marginal band microtubules, suggesting that it may be important for determining microtubule function in vivo.
Highlights
DETERMINATION OF THE EFFECTS OF ERYTHROCYTE TAU, TUBULIN ISOFORMS, AND TUBULIN OLIGOMERS ON ERYTHROCYTETUBULIN ASSEMBLY, AND COMPARISON WITH BRAIN MICROTUBULE ASSEMBLY*
In this paper we compare preparations of microtubule protein from chicken brainanderythrocytes with respect to their properties of nucleation, growth, and critical concentration to determine to what extent the differences in microtubule self-assembly are due to differences in thetubulin variants and toMAPs
The studies presented here suggest that these properties are due to thepresence of a microtubule-associated tau factor and to theunique characteristics of erythrocyte tubulin itself
Summary
DETERMINATION OF THE EFFECTS OF ERYTHROCYTE TAU, TUBULIN ISOFORMS, AND TUBULIN OLIGOMERS ON ERYTHROCYTETUBULIN ASSEMBLY, AND COMPARISON WITH BRAIN MICROTUBULE ASSEMBLY*. Preliminary to exhibit different assemblyproperties in vitro: 1) studies onthe mechanism by whichhigh M , MAPSstimulate erythrocyte tubulin assembleswith greater efficiency assembly have already been reported (9, l l ) , but the details (lower critical concentration, greater elongationrate) regarding tau-promoted assembly remain unclear In this but exhibits a lower nucleatiornatethan brain tubulin, paper, we establish that the50-70-kDa factors inerythrocyte and 2) erythrocytetubulinreadilyformsoligomers microtubule preparations aretau and thathteir primary effect whose presencesignificantly retards therate of elon- is to stimulate microtubule assembly by promoting microtugation, suggesting that tubulin oligomers may be bule nucleation. In this paper we compare the mechanisms of microtubule We have shown that the variants contain biochemically disself-assembly (nucleationand elongation) for two preparations of microtubule protein in the chicken, one from brain tissue, the other from erythrocytes from the blood These preparations exhibit major differences in their self-assembly i n uitro, and have been shown to differ from each other in the kinds of MAPS’ and tubulinisoforms that theycontain [1,2]. The abbreviations used are: MAPs, microtubule-associated protein; SDS, sodium dodecyl sulfate; PC, phosphocellulose; Pipes, 1,4piperazinediethanesulfonicacid
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