2002
DOI: 10.1074/jbc.m110369200 View full text |Buy / Rent full text
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Abstract: We have previously identified the opium alkaloid noscapine as a microtubule interacting agent that binds stoichiometrically to tubulin and alters its conformation. Here we show that, unlike many other microtubule inhibitors, noscapine does not significantly promote or inhibit microtubule polymerization. Instead, it alters the steady-state dynamics of microtubule assembly, primarily by increasing the amount of time that the microtubules spend in an attenuated (pause) state. Further studies reveal that even at h… Show more

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“…Recent work has shown Bub1 and BubR1 levels, but not Mad2 levels, are partially elevated at HeLa cell kinetochores in response to a decrease in tension caused by low levels of vinblastine or noscapine (Skoufias et al, 2001;Zhou et al, 2002). A similar result is found in our PtK1 cells where tension is reduced by cooling to 23°C.…”
Section: Localization Of Kinetochore Components Is Regulated By Attacsupporting
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“…Recent work has shown Bub1 and BubR1 levels, but not Mad2 levels, are partially elevated at HeLa cell kinetochores in response to a decrease in tension caused by low levels of vinblastine or noscapine (Skoufias et al, 2001;Zhou et al, 2002). A similar result is found in our PtK1 cells where tension is reduced by cooling to 23°C.…”
Section: Localization Of Kinetochore Components Is Regulated By Attacsupporting
“…Similar increases in BubR1 levels are seen after treatment of Ptk1 cells with taxol. After loss of tension, Bub1 and BubR1 levels do not increase to prometaphase levels; therefore, microtubule attachment does play a major role in kinetochore localization of these proteins (Hoffman et al, 2001;Skoufias et al, 2001;Zhou et al, 2002). Bub1 levels at kinetochores in HeLa cells are regulated by both attachment and tension, because microtubule attachment to one sister kinetochore causes a decrease in Bub1 only on the attached sister, and lack of tension caused by taxol treatment of metaphase cells increases Bub1 levels at both attached sister kinetochores (Taylor et al, 2001).…”
Section: Localization Of Kinetochore Components Is Regulated By Attacmentioning
“…Multiple attempts have been made in different organisms to uncouple microtubule attachment at the kinetochores from the subsequent development of tension across the kinetochores. Treatment of mammalian cells with microtubule toxins, such as taxol, vinblastine or noscapine at low concentrations or, in the case of PtK cells, exposure to hypothermia, alters microtubule dynamics in such a way that kinetochore microtubule attachment remains largely unaffected, but the tension across the paired kinetochores is absent (Waters et al, 1998;Hoffman et al, 2001;Skoufias et al, 2001;Shannon et al, 2002;Zhou et al, 2002). These cells show a prolonged mitotic arrest under these conditions, suggesting that microtubule attachment per se might not be sufficient to silence the checkpoint.…”
Section: Attachment Versus Tensionmentioning
“…The total cell lysates were collected as described earlier [15,30,31]. The cell lysates containing polymeric and soluble tubulin (monomeric tubulin) were prepared as described earlier [32,33]. Briefly, cells were collected, washed twice in PBS and then incubated at 37 8C with a microtubule stabilization buffer (0.1 M MES, pH 6.75, 1 mM MgSO 4 , 2 mM EGTA, 4 M glycerol) containing 0.1% Triton X-100 for 10 min.…”
Section: Preparation Of Total Cell Lysate and Fractions Containing Pomentioning