Distinct but overlapping sites within the cytoplasmic dynein heavy chain for dimerization and for intermediate chain and light intermediate chain binding

被引:89
作者
Tynan, SH
Gee, MA
Vallee, RB
机构
[1] Univ Massachusetts, Med Ctr, Dept Cell Biol, Worcester, MA 01605 USA
[2] Tropix PE Biosyst, Bedford, MA 01730 USA
关键词
D O I
10.1074/jbc.M001537200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Cytoplasmic dynein is a molecular motor complex consisting of four major classes of polypeptide: the catalytic heavy chains (HC), intermediate chains (IC), light intermediate chains (LIC), and light chains (LC). Previous studies have reported that the ICs bind near the N terminus of the HCs, which is thought to correspond to the base of the dynein complex. In this study, we co-overexpressed cytoplasmic dynein subunits in COS-7 cells to map HC binding sites for the ICs and LICs, as well as HC dimerization. We have found that the LICs bind directly to the N terminus of the HC, adjacent to and overlapping with the IC binding site, consistent with a role for the LICs in cargo binding. Mutation of the LIC P-loop had no detectable effect on RC binding, We detected no direct interaction between the ICs and LICs. Using triple overexpression of HC, IC and LIC, we found that both IC and LIC are present in the same complexes, a result verified by anti-IC immunoprecipitation of endogenous complexes and immunoblotting, Our results indicate that the LICs and ICs must be located on independent surfaces of cytoplasmic dynein to allow each to interact with other proteins without steric interference.
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页码:32769 / 32774
页数:6
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