MAP Kinase Signaling During M Phase Progression 243
tions of cytokinesis in plant and animal cells, the cytoskeletal structures and
themodeofMTdynamicsappeartobelargelyconserved.Notably,however,
plant cytokinesis includes the synthesis of the cell wall in addition to the fu-
sion of cell membranes.
The phenotypes generated by overexpression of the dominant-negative
mutants of NACK1, NPK1, and NQK1 are similar to those of cells treated
with taxol, a compound that blocks the depolymerization of MTs (Yasuhara
et al. 1993). This suggests that MT disassembly is required for phragmoplast
expansion. In cells fromatnack1/hikmutant plants ofArabidopsis, phrag-
moplast MTs persist in the center of the division plane, suggesting that the
disassembly of phragmoplast MTs is inhibited in these cells (Strompen et al.
2002). Phragmoplast MTs usually disappear from the division plane after
formation of the cell plate. Thus, the activation of the NACK-PQR pathway ap-
pears to be required for the reorganization of phragmoplast MTs in expansion
of the phragmoplast during cell plate formation. Factors acting downstream
of the NACK-PQR pathway may therefore control MT dynamics.
3.2
MAP65 Protein is a Downstream Factor of NRK1/NTF6
Identification of NtMAP65-1 as a substrate for NRK1/NTF6 MAPK.Inan-
imals, several components belonging to the KLP or microtubule-associated
protein (MAP) families have been shown to regulate the formation or dy-
namics of the central spindle (Glotzer 2005). In plants, however, few factors
involved in phragmoplast dynamics have been identified.
We found that several MAPs purified from tobacco BY-2 cells can be phos-
phorylated by active NRK1 in vitro. One of these candidate substrates is
NtMAP65-1a, a protein belonging to the MAP65/Ase1/PRC1 family (Sasabe
et al. 2006). This family of proteins is conserved among a variety of or-
ganisms and includes Ase1p (anaphase spindle elongation factor) in yeast
(Pellman et al. 2002), PRC1 (protein regulator of cytokinesis 1) in mam-
mals (Jiang et al. 1998), SPD1 (spindle defective1) inC. elegans (Ver-
brugghe and White 2004), and Feo (Fascetto) inDrosophila(Vernì et al.
2004). Although members of the MAP65/Ase1/PRC1 protein family have
low amino acid sequence similarities, the secondary structures in their pre-
dicted coiled-coil regions are highly conserved (Schuyler et al. 2003). These
MAPs localize to the cytokinetic apparatuses, and most of them are involved
in cytokinesis.
Tobacco has at least three members of the MAP65 subfamily (NtMAP65-
1a, -1b and -1c). The amino acid sequences of these proteins are more than
85 % identical (Hussey et al. 2002), and when produced as recombinant pro-
teins inEscherichia coli, they are effectively phosphorylated by active NRK1
(our unpublished observation). In vitro, NRK1 phosphorylates NtMAP65-1a
at a single site, Thr-579, in the carboxy-terminal region. Specific antibodies