C-terminal domains deliver the DNA replication factor Ciz1 to the nuclear matrix.
Cip1-interacting zinc finger protein 1 (Ciz1) stimulates DNA replication in vitro and is required for mammalian cells to enter S phase. Here, we show that a significant proportion of Ciz1 is retained in nuclear foci following extraction with nuclease and high salt. This suggests that Ciz1 is normally immobilized by interaction with non-chromatin nuclear structures, consistent with the nuclear matrix. Furthermore, matrix-associated Ciz1 foci strikingly colocalize with sites of newly synthesized DNA in S phase nuclei, suggesting that Ciz1 is present in DNA replication factories. Analysis of green fluorescent protein-tagged fragments indicates that nuclear immobilization of Ciz1 is mediated by sequences in its C-terminal third, encoded within amino acids 708-830. Immobilization occurs in a cell-cycle-dependent manner, most probably during late G1 or early S phase, to coincide with its reported point of action. Although C-terminal domains are sufficient for immobilization, N-terminal domains are also required to specify focal organization. Combined with previous work, which showed that the DNA replication activity of Ciz1 is encoded by N-terminal sequences, we suggest that Ciz1 is composed of two functionally distinct domains: an N-terminal replication domain and a C-terminal nuclear matrix anchor. This could contribute to the formation or function of DNA replication factories in mammalian cells.
Ainscough
Justin F-X
JF
CRISTAL, School of Medicine, Leeds University, LS2 9JT, UK.
Rahman
Faisal Abdel
FA
Sercombe
Heather
H
Sedo
Alicia
A
Gerlach
Bjorn
B
Coverley
Dawn
D
D000818:Animals; D004261:DNA Replication / Q000502:physiology*; D003851:Deoxyribonucleases; D049452:Green Fluorescent Proteins / Q000235:genetics; D051379:Mice; D041681:NIH 3T3 Cells; D015530:Nuclear Matrix / Q000235:genetics / Q000378:metabolism*; D009687:Nuclear Proteins / Q000737:chemistry* / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D016196:S Phase / Q000502:physiology; D012492:Salts; D016335:Zinc Fingers / Q000502:physiology*
D009687:Nuclear Proteins; D012492:Salts; D049452:Green Fluorescent Proteins; D003851:Deoxyribonucleases