Targets of the Nackpqr Pathway

The MAP kinase pathway consisting of NPK1, NQK1 and NRK1, which is activated by NACK1, is designated the NACK-PQR pathway (Soyano et al. 2003). The NACK-PQR pathway identified in tobacco seems to be highly conserved in Arabidopsis. Since the MAP kinase pathway itself is a mediator that transmits and propagates the various signals to downstream factors, it is important to identify the targets whose activities are regulated by phospho-rylation by the MAP kinase pathway. In the NACK-PQR pathway, one of the targets of MAPK has been identified as MAP65, a member of the microtubule-associated protein (MAP) family, a gene family that is conserved in eukary-otes, including humans, yeasts and plants in the bundling of microtubules (MTs; Sasabe and Machida 2006). In tobacco cells, NtMAP65-1 is phospho-rylated during mitosis by CDK and NRK1. The phosphorylated NtMAP65-1 is located on the equatorial plane of the phragmoplast whereas unphospho-rylated NtMAP65-1 is more broadly distributed (Sasabe et al. 2006). The overexpression of mutant MAP65-1 with a substitution of the amino acid residue that is phosphorylated by NRK1 results in a delay in cytokinesis. Bundling activity of NtMAP65-1 in vitro is inhibited by phosphorylation by NRK1, and the expression of a non-phosphorylatable form of NtMAP65-1 stabilizes MTs. These results suggest that one of the molecular processes controlled by NRK1 MAP kinase is the regulation of the stability of MTs, and that phosphorylation of NtMAP65-1 by NRK1 MAP kinase contributes to the destabilization of MTs, which might increase the dynamic instability of MTs (depolymerization of MTs and polymerization of tubulins).

The importance of phosphorylation of the MAP65 family protein is also becoming clear from studies with Arabidopsis. The Arabidopsis genome encodes nine members of the MAP65 protein family (Hussey et al. 2002).

AtMAP65-1 has been studied intensively and has been shown to be able to bundle MTs but not to polymerize tubulin (Smertenko et al. 2004, 2006). It is suggested that the bundling activity of AtMAP65-1 is also regulated by phosphorylation, although the responsible kinase has not been identified (Smertenko et al. 2006). This observation is consistent with the results of studies on NtMAP65-1 obtained by Sasabe et al. (2006).

Another member of the Arabidopsis MAP65 family, called AtMAP65-3/ PLEIADE, also has an important role in maintaining the structure of the phragmoplast (Müller et al. 2004). The ple mutant was originally identified as the mutant causing short roots and disorganized cell division (Müller et al. 2002). In the ple mutant root, multinucleated cells and stubs of cell walls were observed, phenotypes resembling those of other cytokinesis mutants. In contrast with other cytokinesis mutants including atnack1 and mkk6, the ple mutant showed abnormal morphology in the roots but not the shoots, suggesting the existence of a homologous gene expressed in the shoots. The lack of biochemical analysis of AtMAP65-3 and mutant analysis of other members of this family prevents us from understanding the roles of this family in plant cell division and development.

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