Abstract
Chromatin Assembly Complex 1 (CAF-1) is a major histone chaperone involved in deposition of histone H3 and H4 into nucleosome. CAF-1 is composed of three subunits; p150, p60 and p48 for human and Cac1, Cac2 and Cac3 for yeast. Despite of its central role in chromatin formation, structural features of the full CAF-1 in complex with histones and other chaperones have not been well characterized. Here, we dissect molecular architecture of yeast CAF-1 (yCAF-1) by cross-linking mass spectrometry (XL-MS) and negative stain single-particle electron microscopy (EM). Our work revealed that Cac1, the largest subunit of yCAF-1, might serve as a major histone binding platform linking Cac2 and Cac3. In addition, EM analysis showed that yCAF-1 adopts a bilobal shape and Cac1 connecting Cac2 and Cac3 to generate a platform for binding histones. This study provides the first structural glimpse of the full CAF-1 complex and a structural framework to understand histone chaperoning processes.
Highlights
Chromatin is the high order structure of eukaryotic DNA and is composed of a repeating unit known as the nucleosome
Of the histone H3/H4 dimer is mediated by Heat shock protein 90 (HSP90), Nuclear Auto-antigen Sperm Protein (NASP), Hat1-RbAp48 histone acetyltransferase and Anti-Silencing Factor-1 (Asf1) in the cytosol
To characterize the structural properties of the fully assembled CAF-1, we developed a baculovirus-expression based procedure to reconstitute the full three-subunit yeast CAF-1 (yCAF-1) (Fig. 1a)
Summary
Chromatin Assembly Factor 1 received: 29 October 2015 accepted: 09 May 2016 Published: 25 May 2016. Chromatin Assembly Complex 1 (CAF-1) is a major histone chaperone involved in deposition of histone H3 and H4 into nucleosome. Despite of its central role in chromatin formation, structural features of the full CAF-1 in complex with histones and other chaperones have not been well characterized. CAF-1 is the major histone chaperone complex for histone H3/H4 and composed of three subunits: p150, p60 and p48 for human, Cac[1], Cac[2] and Cac[3] for yeast, and p180, p105 and p55 for fly[2]. Human p60 and yeast Cac[2] were found to bind to another histone chaperone Asf[1] and this interaction is mediated by histone H3/H49–12. We characterize the molecular architecture of the yeast yCAF-1 complex by combining chemical cross-linking mass spectrometry (XL-MS) and single-particle EM
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