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Figure 5.4 Cytoskeletal alterations produced by active mDia1. (A) Cells expressing
mDia1 DN3 contain an elevated level of actin filaments, as revealed by Cy3-phalloidin
staining. These actin filaments (A) together with microtubules (B,D) align along the axis of
the bipolar cell. As visualized using a GFP-fusion construct of mDia1 DN3 (C) together
with g-tubulin antibody staining (insert in C), the active form of mDia1 localizes to the
centrosome. Bar 10 mm
increase in the fraction of microtubules in which a-tubulin undergoes a specific
post-translational modification, the removal of C-terminal tyrosine by
tubulin-carboxypeptidase (Palazzo et al., 2001). The increased fraction of
the detyrosinated a-tubulin usually correlates with a longer microtubule
lifetime (Khawaja et al., 1988; Webster et al., 1987), so that active mDia might
induce microtubule stabilization. We investigated the effect of mDia1 on
microtubules in more detail, using direct observations of microtubule
dynamics in living cells and measurements of microtubule polymer mass.
Our studies revealed that active mDia1 profoundly affects microtubule
assembly-disassembly processes, both at the 'plus' and 'minus' microtubule
end (Ballestrem et al., 2003; unpublished).
In control cells, expressing normal mDia1, microtubules nucleated at the
centrosome grow toward the cell periphery at a constant speed. When the
microtubule plus end approaches the cell edge, it demonstrates 'dynamic
instability', an oscillatory behaviour with alternating periods of growth and
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