Cell cycle-dependent phosphorylation of human DNA ligase I at the cyclin-dependent kinase sites.
We have described previously that, during S-phase, human DNA ligase I is phosphorylated on Ser66, a casein kinase II site. Here we investigate the phosphorylation status of DNA ligase I during the cell cycle by gel shift analysis and electrospray mass spectrometry. We show that three residues (Ser51, Ser76, and Ser91), which are part of cyclin-dependent kinase sites, are phosphorylated in a cell cycle-dependent manner. Phosphorylation of Ser91 occurs at G1/S transition and depends on a cyclin binding site in the C-terminal part of the protein. This modification is required for the ensuing phosphorylation of Ser76 detectable in G2/M extracts. The substitution of serines at positions 51, 66, 76, and 91 with aspartic acid to mimic the phosphorylated enzyme hampers the association of DNA ligase I with the replication foci. We suggest that the phosphorylation of DNA ligase I and possibly other replicative enzymes is part of the mechanism that directs the disassembly of the replication machinery at the completion of S-phase.
Ferrari
Giovanni
G
Istituto di Genetica Molecolare, Consiglio Nazionale delle Ricerche, via Abbiategrasso 207, 27100 Pavia, Italy.
Rossi
Rossella
R
Arosio
Daniele
D
Vindigni
Alessandro
A
Biamonti
Giuseppe
G
Montecucco
Alessandra
A
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D001665:Binding Sites; D002453:Cell Cycle; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D016213:Cyclins / Q000378:metabolism; D000072481:DNA Ligase ATP; D011088:DNA Ligases / Q000737:chemistry / Q000378:metabolism*; D004261:DNA Replication; D006801:Humans; D008969:Molecular Sequence Data; D010766:Phosphorylation; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism
D016213:Cyclins; C000607892:LIG1 protein, human; D018809:Proliferating Cell Nuclear Antigen; D018844:Cyclin-Dependent Kinases; D011088:DNA Ligases; D000072481:DNA Ligase ATP