Identification of Drosophila Myt1 kinase and its role in Golgi during mitosis

Entry into mitosis is regulated by inhibitory phosphorylation of cdc2/cyclin B, and these phosphorylations can be mediated by the Wee kinase family. Here, we present the identification of Drosophila Mytl (dMytl) kinase and examine the relationship of Mytl and Weel activities in the context of cdc2 phosphorylation. dMytl kinase was found by BLAST-searching the complete Drosophila genome using the amino acid sequence of human Mytl kinase. A single predicted polypeptide was identified that shared a 48% identity, within the kinase domain with human and Xenopus Mytl. Consistent with its putative role as negative regulator of mitotic entry, overexpression of this protein in Drosophila S2 cells resulted in a reduced rate of cellular proliferation while the loss of expression via RNA interference (RNAi) resulted in an increased rate of proliferation. In addition. loss of dMytl alone or in combination with Drosophila Weel (dWeel) resulted in a reduction of cells in G2/M phase and an increase in G1 phase cells. Finally, loss of dMytl alone resulted in a significant reduction of phosphorylation of cdc2 on the threonine-14 (Thr-14) residue as expected. Surprisingly however, a reduction in the phosphorylation of cdc2 on the tyrosine-15 (Tyr-15) residue was only observed when both dMyt1 and dWee1 expression was reduced via RNAi and not by Weel alone. Most strikingly, in the absence of dMytl, Golgi fragmentation during mitosis was incomplete. Our findings suggest that dMytl and dWeel have distinct roles in the regulation of cdc2 phosphorylation and the regulation of mitotic events. (C) 2002 Elsevier Science Inc. All rights reserved.