Asf1 is required for viability and chromatin assembly during DNA replication in vertebrate cells

Asf1 ( anti- silencing function 1), a well conserved protein from yeast to humans, acts as a histone chaperone and is predicted to participate in a variety of chromatin- mediated cellular processes. To investigate the physiological role of vertebrate Asf1 in vivo, we generated a conditional Asf1- deficient mutant from chicken DT40 cells. Induction of Asf1 depletion resulted in the accumulation of cells in S phase with decreased DNA replication and increased mitotic aberrancy forming multipolar spindles, leading to cell death. In addition, nascent chromatin in Asf1- depleted cells showed increased nuclease sensitivity, indicating impaired nucleosome assembly during DNA replication. Complementation analyses revealed that the functional domain of Asf1 for cell viability was confined to the N- terminal core domain ( amino acids 1 - 155) that is a binding platform for histones H3/ H4, CAF- 1p60, and HIRA, whereas Asf1 mutant proteins, abolishing binding abilities with both p60 and HIRA, exhibit no effect on viability. These results together indicate that the vertebrate Asf1 plays a crucial role in replication- coupled chromatin assembly, cell cycle progression, and cellular viability and provide a clue of a possible role in a CAF- 1- and HIRA- independent chromatin- modulating process for cell proliferation.