TY - JOUR

T1 - Repression of DNA replication licensing in quiescence is independent of geminin and may define the cell sycle state of progenitor cells

AU - Kingsbury, Sarah

AU - Loddo, Marco

AU - Fanshawe, Thomas

AU - Obermann, Ellen

AU - Prevost, Toby

AU - Stoeber, Kai

AU - Williams, Gareth

PY - 2005/9/10

Y1 - 2005/9/10

N2 - The DNA replication (or origin) licensing machinery ensures precise duplication of the genome and contributes to the regulation of proliferative capacity in metazoa. Using an in vitro fibroblast model system coupled to a cell-free DNA replication assay, we have studied regulation of the origin licensing pathway during exit from and re-entry into the mitotic cell cycle. We show that in the quiescent state (G0) loss of proliferative capacity is achieved in part through down-regulation of the replication licensing factors Cdc6 and Mcm2–7. The origin licensing repressor geminin is absent in quiescent fibroblasts, suggesting that this powerful inhibitor of the licensing machinery is not required to suppress proliferative capacity in G0. Geminin expression is induced at a late stage in the G0–S transition post pre-RC assembly. Ectopic geminin can block re-acquisition of DNA replication competence during re-entry into the cell cycle, indicating that geminin levels must be tightly down-regulated for escape from G0. Analysis of geminin levels in thyroid shows that geminin expression is suppressed in anatomical compartments/tissues harbouring quiescent cells, confirming our in vitro data. Spatio-temporal control of geminin expression may therefore be of particular relevance for multi-potential stem cells which cycle infrequently. In support of this hypothesis, we have identified a unique population of cells in the putative stem cell niche of intestinal epithelium that are unlicensed and lack geminin expression, a prerequisite for successful re-entry into cycle. Our data argue that the prolonged cell cycle times observed for intestinal stem cells could be due to exit of progenitor cells from cycle into an unlicensed “out-of-cycle” state, a powerful mechanism by which rapidly proliferating tissues may resist genotoxic insult.

AB - The DNA replication (or origin) licensing machinery ensures precise duplication of the genome and contributes to the regulation of proliferative capacity in metazoa. Using an in vitro fibroblast model system coupled to a cell-free DNA replication assay, we have studied regulation of the origin licensing pathway during exit from and re-entry into the mitotic cell cycle. We show that in the quiescent state (G0) loss of proliferative capacity is achieved in part through down-regulation of the replication licensing factors Cdc6 and Mcm2–7. The origin licensing repressor geminin is absent in quiescent fibroblasts, suggesting that this powerful inhibitor of the licensing machinery is not required to suppress proliferative capacity in G0. Geminin expression is induced at a late stage in the G0–S transition post pre-RC assembly. Ectopic geminin can block re-acquisition of DNA replication competence during re-entry into the cell cycle, indicating that geminin levels must be tightly down-regulated for escape from G0. Analysis of geminin levels in thyroid shows that geminin expression is suppressed in anatomical compartments/tissues harbouring quiescent cells, confirming our in vitro data. Spatio-temporal control of geminin expression may therefore be of particular relevance for multi-potential stem cells which cycle infrequently. In support of this hypothesis, we have identified a unique population of cells in the putative stem cell niche of intestinal epithelium that are unlicensed and lack geminin expression, a prerequisite for successful re-entry into cycle. Our data argue that the prolonged cell cycle times observed for intestinal stem cells could be due to exit of progenitor cells from cycle into an unlicensed “out-of-cycle” state, a powerful mechanism by which rapidly proliferating tissues may resist genotoxic insult.

KW - ORC

KW - Cdc6

KW - Mcm2

KW - MCM

KW - Geminin

KW - Pre-RC

KW - Origin licensing

KW - Stem cells

U2 - 10.1016/j.yexcr.2005.05.027

DO - 10.1016/j.yexcr.2005.05.027

M3 - Journal article

VL - 309

SP - 56

EP - 67

JO - Experimental Cell Research

JF - Experimental Cell Research

SN - 0014-4827

IS - 1

ER -