Human geminin promotes pre-RC formation and DNA replication by stabilizing CDT1 in mitosis.
Geminin is an unstable inhibitor of DNA replication that negatively regulates the licensing factor CDT1 and inhibits pre-replicative complex (pre-RC) formation in Xenopus egg extracts. Here we describe a novel function of Geminin. We demonstrate that human Geminin protects CDT1 from proteasome-mediated degradation by inhibiting its ubiquitination. In particular, Geminin ensures basal levels of CDT1 during S phase and its accumulation during mitosis. Consistently, inhibition of Geminin synthesis during M phase leads to impairment of pre-RC formation and DNA replication during the following cell cycle. Moreover, we show that inhibition of CDK1 during mitosis, and not Geminin depletion, is sufficient for premature formation of pre-RCs, indicating that CDK activity is the major mitotic inhibitor of licensing in human cells. Taken together with recent data from our laboratory, our results demonstrate that Geminin is both a negative and positive regulator of pre-RC formation in human cells, playing a positive role in allowing CDT1 accumulation in G2-M, and preventing relicensing of origins in S-G2.
Ballabeni
Andrea
A
Department of Experimental Oncology, European Institute of Oncology, Milan, Italy.
Melixetian
Marina
M
Zamponi
Raffaella
R
Masiero
Laura
L
Marinoni
Federica
F
Helin
Kristian
K
D000818:Animals; D018797:Cell Cycle Proteins / Q000378:metabolism*; D045744:Cell Line, Tumor; D002522:Chlorocebus aethiops; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D004247:DNA / Q000096:biosynthesis / Q000235:genetics; D004261:DNA Replication; D004789:Enzyme Activation; D016193:G1 Phase; D064248:Geminin; D017353:Gene Deletion; D005786:Gene Expression Regulation; D006801:Humans; D008938:Mitosis; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011485:Protein Binding; D014452:Ubiquitins / Q000378:metabolism; D029867:Xenopus Proteins
C467199:CDT1 protein, human; D018797:Cell Cycle Proteins; C577998:GMNN protein, Xenopus; C577995:GMNN protein, human; D064248:Geminin; D014452:Ubiquitins; D029867:Xenopus Proteins; D004247:DNA; D018844:Cyclin-Dependent Kinases; D046988:Proteasome Endopeptidase Complex