Phosphorylation of the C-terminal domain of yeast topoisomerase II by casein kinase II affects DNA-protein interaction.
Eukaryotic DNA topoisomerase II is an abundant nuclear enzyme that is essential for cell proliferation. This homodimeric enzyme catalyzes the cleavage and re-ligation of double-stranded DNA required to separate replicated sister chromatids. Both biochemical and genetic studies show that its catalytic activity is required for chromosome condensation and segregation, and that its decatenation activity can be stimulated by a variety of protein kinases in vitro. In budding yeast, topoisomerase II is most highly phosphorylated in metaphase, and casein kinase II (CKII) was shown to be the major kinase modifying topoisomerase II. We have investigated the effects of phosphorylation of yeast topoisomerase II by CKII in vitro, by means of gel-retardation and filter binding assays. The phosphorylation of the C terminus of topoisomerase II by CKII appears to increase the stability of the complex formed with linear DNA fragments, while dephosphorylation has the opposite effect. Rephosphorylation of phosphatase-treated topoisomerase II by chicken casein kinase II restores a stable protein-DNA complex using a linear DNA fragment. The enhanced stability of the topoisomerase II-DNA complex is also observed with relaxed circular DNA, but not with supercoiled minicircles, in agreement with published results using topoisomerase II from Drosophila. Limited proteolysis and probing with domain-specific antibodies shows that, with the exception of a weakly modified residue between amino acid residues 660 and 1250, all residues modified by casein kinase II are in the last 180 amino acid residues of yeast topoisomerase II.
Dang
Q
Q
Swiss Institute for Experimental Cancer Research (ISREC), Epalinges/Lausanne.
Alghisi
G C
GC
Gasser
S M
SM
D001665:Binding Sites; D047390:Casein Kinase II; D004250:DNA Topoisomerases, Type II / Q000378:metabolism*; D004271:DNA, Fungal / Q000378:metabolism; D004278:DNA, Superhelical / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004587:Electrophoresis, Agar Gel; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics; D012697:Serine Endopeptidases
D004271:DNA, Fungal; D004278:DNA, Superhelical; D004268:DNA-Binding Proteins; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases; D010744:Phosphoric Monoester Hydrolases; D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase; D004250:DNA Topoisomerases, Type II