Most DNA synthesis in HeLa cell nucleus is concentrated in discrete foci. These synthetic sites can be identified by electron microscopy after allowing permeabilized cells to elongate nascent DNA in the presence of biotin-dUTP. Biotin incorporated into nascent DNA can be then immunolabeled with gold particles. Two types of DNA synthetic sites/replication factories can be distinguished at ultrastructural level: (1) electron-dense structures--replication bodies (RB), and (2) focal replication sites with no distinct underlying structure--replication foci (RF). The protein composition of these synthetic sites was studied using double immunogold labeling. We have found that both structures contain (a) proteins involved in DNA replication (DNA polymerase alpha, PCNA), (b) regulators of the cell cycle (cyclin A, cdk2), and (c) RNA processing components like Sm and SS-B/La auto antigens, p80-coilin, hnRNPs A1 and C1/C2. However, at least four regulatory and structural proteins (Cdk1, cyclin B1, PML and lamin B1) differ in their presence in RB and RF. Moreover, in contrast to RF, RB have structural organization. For example, while DNA polymerase alpha, PCNA and hnRNP A1 were diffusely spread throughout RB, hnRNP C1/C2 was found only at the very outside. Surprisingly, RB contained only small amounts of DNA. In conclusion, synthetic sites of both types contain similar but not the same sets of proteins. RB, however, have more developed microarchitecture, apparently with specific functional zones. This data suggest possible differences in genome regions replicated by these two types of replication factories.

Institute of Experimental Medicine Academy of Sciences of the Czech Republic Vídenská 1083 142 20 Prague 4 Krc Czech Republic

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J Cell Biol. 1979 May;81(2):453-7 PubMed

Science. 1992 Sep 18;257(5077):1689-94 PubMed

Oncogene. 2002 Jan 21;21(4):598-610 PubMed

Nucleic Acids Res. 1978 Oct;5(10):3697-714 PubMed

J Mol Biol. 1986 Nov 5;192(1):65-76 PubMed

Mol Cell Biol. 1994 Nov;14(11):7219-25 PubMed

Genes Dev. 1993 Aug;7(8):1559-71 PubMed

J Cell Sci. 1994 Feb;107 ( Pt 2):425-34 PubMed

J Biol Chem. 1997 Sep 26;272(39):24508-13 PubMed

Exp Cell Res. 1994 Dec;215(2):257-62 PubMed

Cell. 1991 Aug 23;66(4):627-35 PubMed

J Cell Biol. 1994 Jun;125(6):1201-12 PubMed

Genes Dev. 2001 Oct 15;15(20):2720-9 PubMed

J Cell Sci. 1988 Jul;90 ( Pt 3):365-78 PubMed

Cell. 1991 Sep 20;66(6):1217-28 PubMed

Annu Rev Biochem. 1998;67:721-51 PubMed

Development. 1989 Feb;105(2):365-78 PubMed

Curr Opin Cell Biol. 1999 Jun;11(3):363-71 PubMed

Semin Cell Dev Biol. 2005 Jun;16(3):311-21 PubMed

J Histochem Cytochem. 1989 Mar;37(3):395-8 PubMed

J Cell Biol. 1997 Nov 3;139(3):579-87 PubMed

J Ultrastruct Res. 1982 Aug;80(2):133-47 PubMed

J Cell Sci. 1995 Nov;108 ( Pt 11):3451-61 PubMed

Nat Cell Biol. 2000 Apr;2(4):232-9 PubMed

Nature. 1988 Apr 7;332(6164):546-8 PubMed

Genes Dev. 1998 Jul 15;12 (14 ):2131-43 PubMed

Exp Cell Res. 1993 May;206(1):43-8 PubMed

Trends Cell Biol. 2002 Oct;12(10):472-8 PubMed

Gene. 2003 May 22;310:1-15 PubMed

Nature. 1995 Mar 23;374(6520):386-8 PubMed

Genes Dev. 1992 Oct;6(10):1874-85 PubMed

J Cell Sci. 1991 Jun;99 ( Pt 2):247-53 PubMed

Oncogene. 1998 Jan 22;16(3):311-20 PubMed

J Biol Chem. 1982 Jul 25;257(14):8391-6 PubMed

J Cell Biol. 1990 Dec;111(6 Pt 1):2247-59 PubMed

J Cell Biol. 1987 Oct;105(4):1549-54 PubMed

J Cell Sci. 1992 Apr;101 ( Pt 4):773-84 PubMed

Trends Cell Biol. 1994 Feb;4(2):48-52 PubMed

J Cell Sci. 1991 Mar;98 ( Pt 3):271-9 PubMed

J Cell Sci. 1993 Jun;105 ( Pt 2):541-50 PubMed

Genes Dev. 2000 Dec 15;14 (24):3102-14 PubMed

J Cell Sci. 2001 May;114(Pt 10):1811-20 PubMed

Bioessays. 1998 Aug;20(8):660-7 PubMed

J Cell Biol. 1992 Oct;119(1):1-15 PubMed

Science. 1968 May 17;160(3829):774-6 PubMed

Cell. 1994 Nov 18;79(4):573-82 PubMed

J Cell Sci Suppl. 1995;19:59-65 PubMed

J Cell Biol. 1991 Oct;115(1):1-17 PubMed

J Cell Biol. 2001 Jul 23;154(2):293-307 PubMed

Annu Rev Biochem. 2002;71:375-403 PubMed

Cell. 1994 Nov 18;79(4):551-5 PubMed

J Cell Sci. 1994 Aug;107 ( Pt 8):2191-202 PubMed

EMBO J. 1990 Sep;9(9):2865-75 PubMed

J Cell Sci. 1992 Mar;101 ( Pt 3):529-45 PubMed

Genes Dev. 2000 Nov 15;14(22):2855-68 PubMed

J Cell Sci. 1995 Feb;108 ( Pt 2):635-44 PubMed

Exp Cell Res. 2004 Aug 1;298(1):58-73 PubMed

Science. 1992 Sep 25;257(5078):1958-61 PubMed

J Cell Sci. 1991 Dec;100 ( Pt 4):869-76 PubMed

Cancer Cell. 2003 Apr;3(4):311-6 PubMed

J Cell Sci. 1992 Nov;103 ( Pt 3):857-62 PubMed

J Cell Sci. 1989 Nov;94 ( Pt 3):471-7 PubMed

J Cell Biol. 1987 Jul;105(1):577-87 PubMed

Dev Cell. 2002 Sep;3(3):329-37 PubMed

Cell. 1992 Nov 27;71(5):865-73 PubMed

J Cell Sci. 2005 Mar 1;118(Pt 5):847-54 PubMed

J Cell Sci. 1997 Oct;110 ( Pt 20):2507-18 PubMed

Oncogene. 2001 Oct 29;20(49):7223-33 PubMed

J Cell Biochem. 1996 Dec 1;63(3):280-91 PubMed

Crit Rev Eukaryot Gene Expr. 2000;10(2):127-33 PubMed

Biochemistry. 1988 Apr 5;27(7):2590-7 PubMed

Nature. 1993 Dec 16;366(6456):701-4 PubMed

J Cell Biol. 1997 Mar 24;136(6):1201-12 PubMed

EMBO J. 2003 Apr 15;22(8):1878-88 PubMed

J Cell Biol. 2000 Apr 17;149(2):271-80 PubMed

Exp Cell Res. 1986 Aug;165(2):291-7 PubMed

EMBO J. 1987 Jul;6(7):2003-10 PubMed

J Struct Biol. 2000 Dec;132(3):201-10 PubMed

J Biol Chem. 2000 May 12;275(19):14509-16 PubMed

Exp Cell Res. 1992 Oct;202(2):211-23 PubMed

Cancer Surv. 1997;29:7-23 PubMed

Cell. 1991 Sep 20;66(6):1197-206 PubMed

J Cell Biol. 1993 Feb;120(3):625-30 PubMed

EMBO J. 1999 Apr 1;18(7):1939-52 PubMed

J Ultrastruct Res. 1967 Aug 30;19(5):474-86 PubMed

J Cell Biol. 1992 Mar;116(5):1095-110 PubMed

Cell. 1993 Apr 23;73(2):361-73 PubMed

Exp Cell Res. 1991 Jul;195(1):27-37 PubMed

Cell. 1997 Sep 19;90(6):1023-9 PubMed

Proc Natl Acad Sci U S A. 1993 Mar 1;90(5):1947-51 PubMed

J Struct Biol. 2004 Dec;148(3):279-89 PubMed

Science. 2003 May 9;300(5621):987-90 PubMed

J Cell Biol. 1984 Dec;99(6):1997-204 PubMed

Cell. 1986 Nov 21;47(4):577-87 PubMed

Exp Cell Res. 1994 Apr;211(2):275-81 PubMed

J Cell Sci. 2005 Jan 1;118(Pt 1):101-12 PubMed

Annu Rev Biochem. 1993;62:289-321 PubMed

Science. 1996 Dec 6;274(5293):1672-7 PubMed

Cytometry. 1992;13(6):603-14 PubMed

Mol Biol Cell. 1997 Jan;8(1):73-82 PubMed

Mol Biol Cell. 1998 Oct;9(10):2987-3001 PubMed

J Cell Biol. 1991 Nov;115(4):919-31 PubMed

Exp Cell Res. 2005 Feb 15;303(2):482-93 PubMed

Genes Dev. 1988 Feb;2(2):215-27 PubMed

Cell. 1992 Jul 10;70(1):127-38 PubMed

Genes Dev. 1993 May;7(5):812-21 PubMed

Mol Cell Biol. 2004 Jul;24(13):5875-86 PubMed

Cell. 1993 Sep 24;74(6):979-92 PubMed

J Exp Med. 1991 Jun 1;173(6):1407-19 PubMed

Cell. 1994 Dec 30;79(7):1233-43 PubMed

J Cell Biol. 2000 Jun 12;149(6):1179-92 PubMed

J Cell Biol. 1989 Jan;108(1):1-11 PubMed

Annu Rev Cell Biol. 1993;9:265-315 PubMed

DNA Replication: From Radioisotopes to Click Chemistry