DAP-like kinase, a member of the death-associated protein kinase family, associates with centrosomes, centromers, and the contractile ring during mitosis.
DAP-like kinase (Dlk) is a nuclear serine/threonine-specific kinase which has been implicated in apoptosis. However, induction of apoptosis by Dlk requires its relocation to the cytoplasm, particularly association with the actin cytoskeleton, which is achieved through interaction with pro-apoptotic protein Par-4. On the other hand, nuclear Dlk does not induce apoptosis and has rather been implicated in transcription. To further explore the biological functions of Dlk, we established a cell clone of MCF-7 cells stably expressing a GFP-Dlk fusion protein at low level. Ectopic expression of GFP-Dlk did not affect the growth properties of the cells. During interphase, GFP-Dlk showed a diffuse nuclear distribution with punctate staining in a subpopulation of cells. During mitosis, however, Dlk was associated with centrosomes, centromeres, and the contractile ring, but not with the mitotic spindle. Association with centrosomes, as confirmed by colocalization with gamma-tubulin and pericentrin persisted throughout mitosis but was also seen in interphase cells. Interestingly, GFP-Dlk and gamma-tubulin could be co-immunoprecipitated indicating that they are present in the same protein complex. Association of Dlk with centromeres, as verified by confocal fluorescence microscopy with centromere-specific antibodies was more restricted and discernable from prophase to early anaphase. Centromere association of Dlk coincides with H3 phosphorylation at Thr11 that is specifically phosphorylated by Dlk in vitro (U. Preuss, G. Landsberg, K. H. Scheidtmann, Nucleic Acids Res. 31, 878-885, 2003). During cytokinesis, Dlk was enriched in the contractile acto-myosin ring and colocalized with Ser19-phosphorylated myosin light chain, which is an in vitro substrate of Dlk. Strikingly, a C-terminal truncation mutant of Dlk generated multi-nucleated cells. Together, these data suggest that Dlk participates in regulation and, perhaps, coordination of mitotis and cytokinesis.
Preuss
Ute
U
Institute of Genetics, University of Bonn, Bonn, Germany. u.preuss@uni-bonn.de
Bierbaum
Hanna
H
Buchenau
Peter
P
Scheidtmann
Karl Heinz
KH
D000199:Actins / Q000378:metabolism; D000818:Animals; D000941:Antigens / Q000378:metabolism; D017209:Apoptosis / Q000502:physiology*; D051017:Apoptosis Regulatory Proteins; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002352:Carrier Proteins / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D002503:Centromere / Q000378:metabolism*; D018385:Centrosome / Q000378:metabolism*; D003599:Cytoskeleton / Q000378:metabolism; D064550:Death-Associated Protein Kinases; D049452:Green Fluorescent Proteins; D006801:Humans; D007399:Interphase / Q000502:physiology; D047908:Intracellular Signaling Peptides and Proteins; D008164:Luminescent Proteins; D020930:MAP Kinase Kinase Kinases; D008856:Microscopy, Fluorescence; D008938:Mitosis / Q000502:physiology; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D051381:Rats; D008941:Spindle Apparatus / Q000378:metabolism*; D014404:Tubulin / Q000378:metabolism
D000199:Actins; D000941:Antigens; D051017:Apoptosis Regulatory Proteins; D002352:Carrier Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008164:Luminescent Proteins; D014404:Tubulin; C086178:pericentrin; C101296:prostate apoptosis response-4 protein; D049452:Green Fluorescent Proteins; D064550:Death-Associated Protein Kinases; D017346:Protein Serine-Threonine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D020930:MAP Kinase Kinase Kinases; C090757:mitogen-activated protein kinase kinase kinase 12