Poster Sessions

Mol-12

Ed Luk

E. Luk, A. Ranjan, G. Mizuguchi, C.-H. Wu, J. Backus, A. Schomburg, W. Wu, C. Wu

Mechanism of histone replacement mediated by the SWR1 complex

The H2A histone variant H2AZ plays roles in various genomic processes. The site-specific incorporation of H2AZ in chromatin is dependent on the activity of the SWR1 chromatin remodeling complex. In vitro, the SWR1 complex replaces H2A-H2B in chromatin with H2AZ-H2B in an ATP-dependent manner. To further dissect this mechanism, we used a photocrosslinking approach to probe interactions between H2AZ-H2B and subunits of the SWR1 complex. Preliminary data showed that the Swc2 and Swr1 subunits physically interact with H2AZ-H2B. Upon ATP hydrolysis, the interaction with Swr1 is lost coincident with the transfer of H2AZ-H2B into the nucleosome. Furthermore, we have employed a native gel assay to measure the number of H2AZ-H2B molecules per nucleosome. Given that a nucleosome has two copies of each histone, we observed the SWR1 complex can replace both copies of H2A-H2B with H2AZ-H2B. Strikingly, nucleosomes containing one H2A-H2B and one H2AZ-H2B are also generated under limiting enzymatic conditions. These heterotypic nucleosomes may represent an intermediate step of the histone replacement reaction. The data is consistent with a model in which the SWR1 complex incorporates H2AZ-H2B into a nucleosome one at a time.