Molecular steps of G-overhang generation at human telomeres and its function in chromosome end protection.
Telomeric G-overhangs are required for the formation of the protective telomere structure and telomerase action. However, the mechanism controlling G-overhang generation at human telomeres is poorly understood. Here, we show that G-overhangs can undergo cell cycle-regulated changes independent of telomerase activity. G-overhangs at lagging telomeres are lengthened in S phase and then shortened in late S/G2 because of C-strand fill-in, whereas the sizes of G-overhangs at leading telomeres remain stable throughout S phase and are lengthened in G2/M. The final nucleotides at measurable C-strands are precisely defined throughout the cell cycle, indicating that C-strand resection is strictly regulated. We demonstrate that C-strand fill-in is mediated by DNA polymerase alpha (polalpha) and controlled by cyclin-dependent kinase 1 (CDK1). Inhibition of CDK1 leads to accumulation of lengthened G-overhangs and induces telomeric DNA damage response. Furthermore, depletion of hStn1 results in elongation of G-overhangs and an increase in telomeric DNA damage. Our results suggest that G-overhang generation at human telomeres is regulated by multiple tightly controlled processes and C-strand fill-in is under the control of polalpha and CDK1.
Dai
Xueyu
X
WWAMI Medical Education Program, Washington State University, Spokane, WA 99210-1495, USA.
Huang
Chenhui
C
Bhusari
Amruta
A
Sampathi
Shilpa
S
Schubert
Kathryn
K
Chai
Weihang
W
D016203:CDC2 Protein Kinase / Q000037:antagonists & inhibitors / Q000378:metabolism; D002453:Cell Cycle; D002460:Cell Line; D004249:DNA Damage; D006367:HeLa Cells; D006801:Humans; D009711:Nucleotides / Q000378:metabolism; D019098:Telomerase / Q000378:metabolism*; D016615:Telomere / Q000737:chemistry / Q000378:metabolism*; D034501:Telomere-Binding Proteins / Q000378:metabolism
D009711:Nucleotides; C544946:Stn1 protein, human; D034501:Telomere-Binding Proteins; D016203:CDC2 Protein Kinase; D019098:Telomerase