ANTIBODIES TO THE KINESIN MOTOR DOMAIN AND CENP-E INHIBIT MICROTUBULE DEPOLYMERIZATION-DEPENDENT MOTION OF CHROMOSOMES IN-VITRO

被引:190
作者
LOMBILLO, VA
NISLOW, C
YEN, TJ
GELFAND, VI
MCINTOSH, JR
机构
[1] UNIV COLORADO,DEPT MOLEC CELLULAR & DEV BIOL,BOULDER,CO 80309
[2] FOX CHASE CANC CTR,PHILADELPHIA,PA 19111
[3] UNIV ILLINOIS,DEPT CELL & STRUCT BIOL,URBANA,IL 61801
关键词
D O I
10.1083/jcb.128.1.107
中图分类号
Q2 [细胞生物学];
学科分类号
071009 ; 090102 ;
摘要
Chromosomes can move with the ends of depolymerizing microtubules (MTs) in vitro, even in the absence of nucleotide triphosphates (Coue, M., V. A. Lombillo, and J. R. McIntosh. 1991. J. Cell Biol. 112:1165-1175.) Here, we describe an immunological investigation of the proteins important for this form of motility. Affinity-purified polyclonal antibodies to kinesin exert a severe inhibitory effect on depolymerization-dependent chromosome motion. These antibodies predominantly recognize a polypeptide of M(r) similar to 250 kD on immunoblots of CHO chromosomes and stain kinetochores as well as some vesicles that are in the chromosome preparation. Antibodies to CENP-E, a kinetochore-associated kinesin-like protein, also recognize a 250-kD electrophoretic component, but they stain only the kinetochroe region of isolated chromosomes. Polyclonal antibodies that recognize specific domains of the CENP-E polypeptide affect MT disassembly-dependent chromosome motion in different ways; antibodies to the head or tail portions slow motility threefold, while those raised against the neck region stop motion completely. Analogous antibodies that block conventional, ATP-dependent motility of cytoplasmic dynein (Vaisberg, G., M. P. Koonce, and J. R. McIntosh. 1993. J. Cell Biol. 123:849-858) have no effect on disassembly-dependent chromosome motion, even though they bind to kinetochores. These observations suggest that CENP-E helps couple chromosomes to depolymerizing MTs. A similar coupling activity may allow spindle MTs to remain kinetochore-bound while their lengths change during both prometaphase and anaphase A.
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页码:107 / 115
页数:9
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