CAF-1 p150 and Ki-67 Regulate Nuclear Structure Throughout the Human Cell Cycle

Abstract

The three-dimensional organization of the human genome is non-random in interphase cells. Heterochromatin is highly clustered at the nuclear periphery, adjacent to nucleoli, and near centromeres. These localizations are reshuffled during mitosis when the chromosomes are condensed, nucleoli disassembled, and the nuclear envelope broken down. After cytokinesis, heterochromatin is re-localized to the domains described above. However, the mechanisms by which this localization is coordinated are not well understood. This dissertation will present evidence showing that both CAF-1 p150 and Ki-67 regulate nuclear structure throughout the human cell cycle. Chromatin Assembly Factor 1 (CAF-1) is a highly conserved three-subunit protein complex which deposits histones (H3/H4) 2 heterotetramers onto replicating DNA during S-phase of the cell cycle. The N-terminal domain of the largest subunit of CAF-1 (p150N) is dispensable for histone deposition, and instead regulates the localization of specific loci (Nucleolar-Associated Domains, or “NADs”) and several proteins to the nucleolus during interphase. One of the proteins regulated by p150N is Ki-67, a protein widely used as a clinical marker of cellular proliferation. Depletion of Ki-67 decreases the association of NADs to the nucleolus in a manner similar to that of p150. Ki-67 is also a fundamental component of the perichromosomal layer (PCL), a sheath of proteins that surrounds all condensed chromosomes during mitosis. A subset of p150 localizes to the PCL during mitosis, and depletion of p150 disrupts Ki-67 localization to the PCL. This activity was mapped to the Sumoylation Interacting Motif (SIM) within p150N, which is also required for the localization of NADs and Ki-67 to the nucleolus during interphase. Together, these studies indicate that p150N coordinates the three-dimensional arrangement of both interphase and mitotic chromosomes via Ki-67.