Journal Article

The Cac1 subunit of histone chaperone CAF-1 organizes CAF-1-H3/H4 architecture and tetramerizes histones

Ahn, Natalie G, Shen, Zih-Jie, Tyler, Jessica K, Nemkov, Travis, Roemer, Sarah C, Churchill, Mair EA, Hansen, Kirk C, Zhou, Yeyun, Liu, Wallace H, Liddle, Jennifer C, Dennehey, Briana K, Balsbaugh, Jeremy L

2016

Overview

The histone chaperone Chromatin Assembly Factor 1 (CAF-1) deposits tetrameric (H3/H4)2 histones onto newly-synthesized DNA during DNA replication. To understand the mechanism of the tri-subunit CAF-1 complex in this process, we investigated the protein-protein interactions within the CAF-1-H3/H4 architecture using biophysical and biochemical approaches. Hydrogen/deuterium exchange and chemical cross-linking coupled to mass spectrometry reveal interactions that are essential for CAF-1 function in budding yeast, and importantly indicate that the Cac1 subunit functions as a scaffold within the CAF-1-H3/H4 complex. Cac1 alone not only binds H3/H4 with high affinity, but also promotes histone tetramerization independent of the other subunits. Moreover, we identify a minimal region in the C-terminus of Cac1, including the structured winged helix domain and glutamate/aspartate-rich domain, which is sufficient to induce (H3/H4)2 tetramerization. These findings reveal a key role of Cac1 in histone tetramerization, providing a new model for CAF-1-H3/H4 architecture and function during eukaryotic replication. The DNA of a human, yeast or other eukaryotic cell is bound to proteins called histones to form repeating units called nucleosomes. Every time a eukaryotic cell divides, it must duplicate its DNA. Old histones are first removed from the nucleosomes before being re-assembled onto the newly duplicated DNA along with new histone proteins, producing a full complement of nucleosomes. A group of proteins called the chromatin assembly factor 1 (or CAF-1 for short) helps to assemble the histones onto the DNA. CAF-1 is made up of three proteins, and binds to two copies of each of the histones known as H3 and H4. These are the first histones to be assembled onto the nucleosomes. It was not clear how the components of CAF-1 are organized, or how CAF-1 recognizes histones. Liu et al. have now investigated the structure of CAF-1 and its interactions with the H3 and H4 histones by studying yeast proteins and cells. Yeast is a good model system because yeast CAF-1 is smaller and easier to isolate than human CAF-1, yet still performs the same essential activities. Using a combination of biochemical and biophysical techniques, Liu et al. found that one of the three proteins that makes up yeast CAF-1 – called Cac1 – forms a scaffold that supports the other CAF-1 proteins and histones H3 and H4. Moreover, a specific part of Cac1 is able to bind to these histones and assemble two copies of each of them to prepare for efficient nucleosome assembly. Further experiments revealed the specific areas where the CAF-1 proteins interact with each other and with the histones, determined how strong those interactions are, and confirmed that these interactions play important roles in yeast. Overall, the results presented by Liu et al. provide new insights into the structure of CAF-1 bound to H3 and H4. In order to understand in detail how CAF-1 helps to assemble histones onto DNA, future work needs to capture three-dimensional snapshots of the different steps in this process. Further investigation is also needed to discover how CAF-1 cooperates with other factors that promote DNA duplication.

• Publisher: eLife Science Publications, Ltd,eLife Sciences Publications Ltd,eLife Sciences Publications, Ltd
• Subject: Architecture / Biochemistry / Biophysics and Structural Biology / C-Terminus / Chromatin remodeling / Deoxyribonucleic acid / DNA / DNA biosynthesis / Experiments / Genes and Chromosomes / histone chaperone / Histones / Laboratories / mass spectrometry / Mass spectroscopy / Medicine / Molecular chaperones / Nucleosome / Observations / Peptides / Physiological aspects / Protein interaction / Protein-protein interactions / Proteins / Replication / structural biology