1999 Fiscal Year Final Research Report Summary
A study on MTs in the cell cortex by HP freezing
| Project/Area Number |
09044225
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| Research Category |
Grant-in-Aid for Scientific Research (B).
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
生物形態・構造
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| Research Institution | Hiroshima University |
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Principal Investigator |
MINEYUKI Yoshinobu HIROSHIMA UNIVERSITY, FACULTY OF SCIENCE, ASSOCIATE PROFESSOR, 理学部, 助教授 (30219703)
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| Co-Investigator(Kenkyū-buntansha) |
KARAHARA Ichirou TOYAMA UNIVERSITY, FACULTY OF SCIENCE, RESEARCH ASSOCIATE, 理学部, 助手 (60283058)
MURATA Takashi UNIVERSITY OF TOKYO, GRADUATE SCHOOL OF ARTS AND SCIENCE, ASSISTANT PROFESSOR, 大学院・総合文化研究科, 講師 (00242024)
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| Project Period (FY) |
1997 – 1999
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| Keywords | High pressure freezing / Electron microscope / Microtubules / preprophase band / Cytokinesis / Coated vesicle / Root meristems / Tomography |
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| Research Abstract |
Interphase cortical microtubules (MTs) and the preprophase band (PPB) are two cortical arrays of MTs in higher plants, and are known to play ezzentrrial role for the regulation of the direction of cell elongation and for the determination of the division site, respectively. However, little is known about the mechanism that regulates these cortical MT organization and turnover. We have employed a combination of high pressure freezing and freeze-substitution or freeze-etch techniques to preserve and visualize forming MTs in onion and tobacco seedlings for ultrastructural analysis. These studies have led to the detection of several novel structural features that have not been reported previously.
(1) In vitro studies of polymerizing/depolymerizing MTs have shown that their ends display both flared and non-flared configurations. Our micrographs demonstrate that cortical MTs also exhibit flared and non-flared ends, consistent with the idea that they are turning over. The number of coiled end configurations (expected depolymerizing MT end in vivo) increased by the treatment with a MT-depolymerizing drug, oryzalin. This suggests that the the frequency of coiled ends reflects that of shrinklng MTs in living cells. (2) Drugs that affect F-actin stability have been shown to perturb PPB morphology. To learn more about how such drugs may cause these changes, we are analyzing quantitatively their effects on both MT-MT and MT-plasma membrane spacings. This type of analysis is more easily done with cryofixed than with chemically fixed samples due to the smooth, non-wavy conformation of the plasma membrane in high pressure frozen cells. (3) Previous studies have shown that PPB regions contain vesicles that stain similar to cell walls, which has led to the suggestion that they could be Involved in delivering special types of cell wall molecules to PPB regions. Our micrographs demonstrate, however, that increased numbers of coated, endocytic vesicles are formed between the PPB MTs.
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