MCM3-binding GANP DNA-primase is associated with a novel phosphatase component G5PR.
BACKGROUND
GANP, carrying DNA-primase and MCM3-binding domains, is up-regulated in germinal centre B cells. To understand the regulatory function of GANP upon MCM complex, we searched for GANP-associated molecules by yeast two-hybrid screening.


RESULTS
Using the 1 kb fragment (G5) of the ganp cDNA, we identified a clone named G5PR that is structurally homologous to known regulatory subunits of protein phosphatases (PPases) and determined the association of G5PR with GANP in vivo in the DNA transfectant. G5PR is associated with protein phosphatase 5 (PP5) through its tetratricopeptide-repeat (TPR) domain. Pull-down assays demonstrated that G5PR is also associated with protein phosphatase 2A (PP2A), the complex of A subunit (PR65) and the catalytic (C) subunit (PP2Ac), similar to the B" subunit. The G5PR-associated complex had phosphatase activity on casein, histone H1 and MCM3 in vitro, but the addition of G5PR did not stimulate or inhibit the phosphatase activities of PP5 and PP2A. The cellular localization of G5PR in transfected cells varies during cell cycling, appearing in the nucleus during prophase, in the peri-chromatin during mitotic phase, and in the cytoplasm after cell division.


CONCLUSIONS
G5PR is capable of recruiting two kinds of PPases, PP5 and PP2A, into the GANP/MCM3 complex, which might regulate its phosphorylation state during cell cycle progression.
Kono
Yoshihiko
Y
Department of Immunology, Kumamoto University School of Medicine, 2-2-1, Honjo, Kumamoto, 860-0811, Japan.
Maeda
Kazuhiko
K
Kuwahara
Kazuhiko
K
Yamamoto
Hideyuki
H
Miyamoto
Eishichi
E
Yonezawa
Kazuyoshi
K
Takagi
Katsumasa
K
Sakaguchi
Nobuo
N
D000123:Acetyltransferases / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D018797:Cell Cycle Proteins; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D064126:Minichromosome Maintenance Complex Component 3; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism*; D010749:Phosphoprotein Phosphatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism*; D054648:Protein Phosphatase 2; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D014157:Transcription Factors
D018797:Cell Cycle Proteins; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D047908:Intracellular Signaling Peptides and Proteins; C066470:MCM3 protein, human; C513269:Mcm3 protein, mouse; D009687:Nuclear Proteins; D010750:Phosphoproteins; D014157:Transcription Factors; D000123:Acetyltransferases; C406825:MCM3AP protein, human; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2; D064126:Minichromosome Maintenance Complex Component 3