Proteolytic analysis of polymerized maize tubulin: Regulation of microtubule stability to low temperature and Ca2+ by the carboxyl terminus of beta-tubulin

被引:20
作者
Bokros, CL [1 ]
Hugdahl, JD [1 ]
Blumenthal, SSD [1 ]
Morejohn, LC [1 ]
机构
[1] UNIV TEXAS, DEPT BOT, AUSTIN, TX 78713 USA
关键词
Zea mays L; chilling injury; low temperature; maize; microtubules; proteolytic analysis; subtilisin; taxol; tubulin;
D O I
10.1111/j.1365-3040.1996.tb00387.x
中图分类号
Q94 [植物学];
学科分类号
071001 ;
摘要
To obtain information on plant microtubule stability to low temperature and Ca2(+), the regulatory domain of polymerized tubulin from maize (Zea mays cv. Black Mexican Sweet) was dissected by limited proteolysis with subtilisin. Tubulin in taxol-stabilized microtubules was cleaved in a subtilisin concentration- and time-dependent manner. Immunoblotting of microtubules with antibodies having mapped epitopes on alpha- and beta-tubulins revealed that cleavage initially removed less than or equal to 15 residues from the beta-tubulin carboxyl terminus to produce alpha beta(s)-microtubules. Subsequent cleavage occurred at an extreme site and an internal site within the alpha-tubulin carboxyl terminus. Electron microscopy revealed that alpha beta(s)-microtubules were ultrastructurally indistinguishable from uncleaved control alpha beta-microtubules. Quantitative polymer sedimentation showed that low temperature treatment (0 degrees C) caused significant depolymerization of alpha beta-microtubules, but little depolymerization of alpha beta(s)-microtubules. Ca2+ enhanced the cold-induced depolymerization of both alpha beta- and alpha beta(s)-microtubules. However, alpha beta(s)-microtubules were significantly more stable to depolymerization by cold and Ca2+ than were alpha beta-microtubules. The results showed that maize microtubules containing shortened beta-tubulin carboxyl termini are relatively resistant to the combined depolymerizing effects of cold and Ca2+. Thus, the extreme carboxyl terminus of beta-tubulin is a crucial element of the plant tubulin regulatory domain and may be involved in the modulation of microtubule stability during the chilling response in plants.
引用
收藏
页码:539 / 548
页数:10
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