MAP Kinase Signaling During M Phase Progression 241
The requirement of NQK1 for the expansion of the phragmoplast and for
the formation of cell plates was demonstrated by overexpressing a kinase-
defective mutant of NQK1 (NQK1:KW) in BY-2 cells and tobacco plants. The
cells overexpressing NQK1:KW are multinucleated and form incomplete cell
plates (Soyano et al. 2003). Also, the seedlings of the NQK1:KW transgenic
plants are poorly developed, and its surfaces have bi- or un-nucleated guard
and pavement cells, similar to those of plants overexpressing NACK:ST. We
observed consistent abnormalities in the cells in which the kinase-defective
NQK1 was overexpressed; therefore, NQK1 MAPKK seems to promote pro-
gression of cytokinesis downstream of NPK1 MAPKKK.
This idea is further supported by observation of the mutant ofANQ1/
AtMKK6,whichistheArabidopsishomolog ofNQK1.Liketheatnack1mutant
ofArabidopsis,theanq1homozygous mutant ofArabidopsisexhibits a severe
dwarf phenotype (Fig. 4B; Soyano et al. 2003). Theanq1mutants have typi-
cal defects in cytokinesis, large cells with incomplete cell walls, and multiple
nuclei in leaves and roots. In addition to the growth defect,anq1mutants
produce abnormal pollen grains that exhibit a tetrad structure (Soyano et al.
2003). This phenotype of pollen grains is similar to those fromtetraspore,
stud,andatnack2mutants, which have allelic mutations (Hülskamp et al.
1997; Spielman et al. 1997; Tanaka et al. 2004) and cause defects in male mei-
otic cytokinesis. Thus, ANQ1/AtMKK6 may also be involved in mitotic and
meiotic cytokinesis under the regulation of NACK.
IdentificationofNRK1MAPKthatactsdownstreamofNQK1MAPKK.
NRK1 MAPK, which may act downstream of NQK1, was identified by yeast
two-hybrid screening using NQK1:KW as the bait (Soyano et al. 2003). The
predicted amino acid sequence of NRK1 is almost identical to that of NTF6
(Calderini et al. 2001). We found that recombinant NRK1 and NQK1 pro-
teins bind each other in vitro (Soyano et al. 2003), which is consistent with
a report by Calderini et al. (2001). Active recombinant NQK1 MAPKK phos-
phorylates and activates NRK1 MAPK (Soyano et al. 2003). Calderini et al.
(1998) reported that NTF6 localizes to the midzone of phragmoplasts in
anaphase tobacco cells. Using NRK1-specific antibodies, we also recently
found that NRK1 localizes to the equator of phragmoplasts at telophase
(our unpublished observation). Although NRK1 protein is present through-
out cell cycle, the activity of NRK1 is activated in parallel with NQK1 at
late M phase (Fig. 3; Soyano et al. 2003), suggesting that NQK1 phosphory-
lates NRK1 in vivo. We designated the kinase cascade that includes NPK1,
NQK1, NRK1, NACK1, and NACK2, the NACK-PQR pathway (Fig. 1; Soyano
et al. 2003).
Although the activation of NRK1 is tightly coupled to the activation of
NPK1 and NQK1, involvement of NRK1 in cytokinesis has not yet been ex-
perimentally demonstrated. The preliminary observations in our laboratory
indicate that the loss-of-function mutants of some MAPKs in Group B inAra-
bidopsisare defective in cytokinesis (our unpublished observations). These