Experimental manipulation of γ‐tubulin distribution in arabidopsis using anti‐microtubule drugs

Abstract

gamma-Tubulin-specific antibodies stain the microtubule (Mt) arrays of Arabidopsis suspension cells in a punctate or patchy manner. During division, staining of kinetochore fibers and the phragmoplast is extensive, except in the vicinity of the plus ends at the metaphase plate and cell plate. gamma-Tubulin localization responds to low levels of colchicine, with staining receding farther toward the minus (pole) ends of kinetochore fibers. At higher drug concentrations, gamma-tubulin also associates with abnormal Mt foci as well as with the surface of the daughter nuclei facing the phragmoplast. During UV-induced recovery from colchicine, gamma-tubulin increases along the presumptive minus ends of mitotic Mts as well as the phragmoplast near the daughter nuclei. With CIPC, immunostaining is concentrated around the centers of focal Mt arrays in multipolar spindles. In the presence of taxol, Mts are more prominent but the mitotic apparatus and phragmoplast are abnormal. As with CIPC, gamma-tubulin is concentrated at focal arrays. Increased punctate staining is also present in interphase arrays, with fluorescent dots often located at the ends of Mts. These results support a preferential association between gamma-tubulin and Mt minus ends, but are also consistent with more general binding along the walls of Mts. Thus, minus ends (and Mt nucleation sites) may be present throughout plant Mt arrays, but gamma-tubulin may also serve another function, such as in structural stabilization.

Interaction of Antiparallel Microtubules in the Phragmoplast Is Mediated by the Microtubule-Associated Protein MAP65-3 inArabidopsis

“…This observation contradicts the conventional view of how antiparallel MTs are held together in the phragmoplast. However, it is in agreement with immunofluorescence images of antitubulin and images of green fluorescent protein (GFP)-marked MTs in somatic cells, because the corresponding images show a dark line in the middle of the phragmoplast (Liu et al, 1995;Yoneda et al, 2005).…”

The γ -Tubulin Complex Protein GCP4 Is Required for Organizing Functional Microtubule Arrays inArabidopsis thaliana

“…Instead, its signal appears as a gradient with more prominent signal toward the spindle poles and the edges of the phragmoplast (Liu et al, , 1994(Liu et al, , 1995. Such a distribution pattern was later confirmed in animal cells (Lajoie-Mazenc et al, 1994;Lü ders and Stearns, 2007).…”

“…In the mitotic spindle and the phragmoplast, g-tubulin appears along MTs but preferentially decorates MT minus ends toward spindle poles and the edges of phragmoplast MTs facing daughter nuclei (Liu et al, , 1995. The localization of g-tubulin along MTs has been verified in spindles and midbodies in animal cells (Lajoie-Mazenc et al, 1994;Lü ders and Stearns, 2007).…”

Meiotic telomere clustering is inhibited by colchicine but does not require cytoplasmic microtubules

“…Colchicine is an effective MT depolymerizing agent in plants at millimolar or greater concentrations (Morejohn, 1991), while at 100 µM, its effects on MTs are less pronounced or undetectable (Liu et al, 1995). Previous experiments examining the effect of colchicine on meiotic prophase have used high colchicine concentrations (1-25 mM), which have the potential of toxic side effects.…”