Cell type-specific responses of human cells to inhibition of replication licensing.
Replication origins are 'licensed' for a single initiation event by loading Mcm2-7 complexes during late mitosis and G1. Licensing is blocked at other cell cycle stages by the activity of cyclin-dependent kinases and a small protein called geminin. Here, we describe the effects of over-expressing a non-degradable form of geminin in various cell lines. Geminin expression reduced the quantity of Mcm2 bound to chromatin and blocked cell proliferation. U2OS (p53+/Rb+) cells showed an early S phase arrest with high cyclin E and undetectable cyclin A levels, consistent with the activation of an intra-S checkpoint. Saos2 (p53-/Rb-) cells showed an accumulation of cells in late S and G2/M with approximately normal levels of cyclin A, consistent with loss of this intra-S phase checkpoint. Geminin also induced apoptosis in both these cell lines. In contrast, IMR90 primary fibroblasts over-expressing geminin arrested in G1 with reduced cyclin E levels and no detectable apoptosis. A 'licensing checkpoint' may therefore act in primary cells to prevent passage into S phase in the absence of sufficient origin licensing. These results suggest that inhibition of the licensing system may cause cancer-specific cell killing and therefore represent a novel anti-cancer target.
Shreeram
S
S
Wellcome Trust Biocentre, University of Dundee, Dow Street, Dundee DD1 5EH, UK.
Sparks
Alison
A
Lane
David P
DP
Blow
J Julian
JJ
D000256:Adenoviridae / Q000235:genetics; D000970:Antineoplastic Agents / Q000494:pharmacology; D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000502:physiology*; D002455:Cell Division; D004261:DNA Replication / Q000187:drug effects; D004268:DNA-Binding Proteins / Q000502:physiology; D064248:Geminin; D006801:Humans; D064111:Minichromosome Maintenance Complex Component 2; D064168:Minichromosome Maintenance Complex Component 7; D009687:Nuclear Proteins / Q000502:physiology; D014407:Tumor Cells, Cultured
D000970:Antineoplastic Agents; C467199:CDT1 protein, human; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C577995:GMNN protein, human; D064248:Geminin; D009687:Nuclear Proteins; C101624:MCM7 protein, human; D064111:Minichromosome Maintenance Complex Component 2; D064168:Minichromosome Maintenance Complex Component 7