MOLECULAR-CLONING AND CHARACTERIZATION OF MAIZE ZMMEK1, A PROTEIN-KINASE WITH A CATALYTIC DOMAIN HOMOLOGOUS TO MITOGEN-ACTIVATED AND STRESS-ACTIVATED PROTEIN-KINASE KINASES

Mitogen- or stress-activated protein kinase kinases (M/SAPKKs) are dua l-specificity protein kinases that are components of highly conserved signal transduction pathways. A cDNA clone (ZmMEK1) was isolated from a Zea mays (L.) root tip library. ZmMEK1 contains a complete open read ing frame, encoding a 355-amino-acid protein with an estimated molecul ar mass of 39,874 Da. The predicted protein contains the 11 catalytic sub-domains conserved in all protein kinases, and a version of the sub -domain VIII S/TxxxS/TxVCJT motif that is characteristic of M/SAPKK pr oteins (EC 2.7.1.37). The catalytic domain of ZmMEK1 is most closely r elated (65% identity) to the tomato M/SAPKK homolog LeMEK1, but exhibi ts similar identity (39-60%) to M/SAPKKs from other plants, animals an d fungi. Northern blotting revealed ZmMEK1 mRNA in maize seedling root s and coleoptiles; in mature plants transcripts were detected in stems and low levels in leaves. Transcription of ZmMEK1 mRNA was also affec ted by environmental stimuli. The catalytic domain of ZmMEK1 was expre ssed as a glutathione-S-transferase (GST) fusion protein in Escherichi a coli. Nanogram quantities of the purified fusion protein reacted wit h anti-M/SAPKK antibodies on immunoblots. In vitro, the GST-ZmMEK1 fus ion protein undergoes autophosphorylation, and will phosphorylate myel in basic protein, but will not phosphorylate histone H1. ZmMEK1 encode s an enzyme that is structurally and functionally similar to other M/S APKK proteins.