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Molecular and Cellular Biology, June 2008, p. 3672-3685, Vol. 28, No. 11
0270-7306/08/$08.00+0     doi:10.1128/MCB.00510-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

In Vivo Study of the Nucleosome Assembly Functions of ASF1 Histone Chaperones in Human Cells{triangledown} ,{dagger}

Angélique Galvani,{ddagger},§ Régis Courbeyrette,§ Morgane Agez, Françoise Ochsenbein, Carl Mann,* and Jean-Yves Thuret*

CEA, iBiTec-S, Gif-sur-Yvette F-91191, France

Received 23 March 2007/ Returned for modification 24 May 2007/ Accepted 20 March 2008

Histone chaperones have been implicated in nucleosome assembly and disassembly as well as histone modification. ASF1 is a highly conserved histone H3/H4 chaperone that synergizes in vitro with two other histone chaperones, chromatin assembly factor 1 (CAF-1) and histone repression A factor (HIRA), in DNA synthesis-coupled and DNA synthesis-independent nucleosome assembly. Here, we identify mutants of histones H3.1 and H3.3 that are unable to interact with human ASF1A and ASF1B isoforms but that are still competent to bind CAF-1 and HIRA, respectively. We show that these mutant histones are inefficiently deposited into chromatin in vivo. Furthermore, we found that both ASF1A and ASF1B participate in the DNA synthesis-independent deposition of H3.3 in HeLa cells, thus highlighting an unexpected role for ASF1B in this pathway. This pathway does not require interaction of ASF1 with HIRA. We provide the first direct determination that ASF1A and ASF1B play a role in the efficiency of nucleosome assembly in vivo in human cells.

* Corresponding author. Mailing address: CEA, SBIGeM-Bât. 142, 91191 Gif-sur-Yvette Cedex, France. Phone for Jean-Yves Thuret: 33 1 69 08 64 25. Fax: 33 1 69 08 80 46. E-mail: jean-yves.thuret{at}cea.fr. Phone for Carl Mann: 33 1 69 08 92 94. Fax: 33 1 69 08 80 46. E-mail: carl.mann{at}cea.fr

{triangledown} Published ahead of print on 31 March 2008.

{dagger} Supplemental material for this article may be found at http://mcb.asm.org/.

{ddagger} Present address: INSERM and Université Denis Diderot-Paris7, UMR S 741, Paris F-75005, France.

§ These two authors contributed equally to this work.

Molecular and Cellular Biology, June 2008, p. 3672-3685, Vol. 28, No. 11
0270-7306/08/$08.00+0     doi:10.1128/MCB.00510-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.





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