Perichromosomal protein Ki67 supports mitotic chromosome architecture

Abstract

Although the condensin complexes and topoisomerase IIa (TopoIIa) are the central players in mitotic chromosome formation, they are insufficient for its completion, and additional factors involved in the process have been extensively sought. In this study, we examined the possibility that Ki67, a perichromosomal protein widely used as a cell proliferation marker, is one such factor. Using a combination of auxin-inducible degron and CRISPR-Cas9-based gene editing technologies, we generated a human HCT116 cell line in which Ki67 is rapidly depleted in a few hours. The removal of Ki67 before mitotic entry did not impact the early mitotic chromosome assembly observed in prophase but subsequently resulted in the formation of misshapen mitotic chromosomes. When Ki67 was removed after mitotic entry, preassembled rod-shaped mitotic chromosomes became disorganized. In addition, we show that Ki67 and TopoIIa are reciprocally coimmunoprecipitated from mitotic cell extracts. These observations indicate that Ki67 aids the finalization of mitotic chromosome formation and helps maintain rod-shaped chromosome architecture, likely in collaboration with TopoIIa. Together, these findings represent a new model in which mitotic chromosome architecture is supported both internally and externally.

<b>Quantitative analyses of the metaphase-to-anaphase transition reveal differential kinetic regulation for securin and cyclin </b><b>B1 </b>

“…5A, B), as in previous study (27). Among potential cdk1 substrates that are being dephosphorylated at the onset of anaphase, we focused to Ki67 as it is characterized as an abundant chromosomal protein which has been implicated in mitotic chromosome architecture (1,24,25,28). Interestingly Ki67 is found to confer an electrostatic charge barrier that assists individualization of chromatids (5).…”

Ki-67 and the Chromosome Periphery Compartment in Mitosis

“…This phenotype was only observed following nuclear envelope breakdown at prometaphase. Further experiments revealed that Ki-67 depletion resulted in the mislocalisation of both Topo IIα and condensin II complex member hCAP-H2, suggesting collaboration between chromosome periphery proteins and chromosome structure proteins [52].…”

“…Some of the best-known periphery components include the nucleolar proteins fibrillarin, nucleolin, nucleophosmin, peripherin and Ki-67 [25,38,43,[46][47][48]. Although the role of most of these proteins at the mitotic chromosome periphery has received little attention, Ki-67 has recently emerged as a particularly active subject for studies [39,[49][50][51][52][53].…”

“…To explore the function(s) of Ki-67 in interphase and mitosis, a Ki-67-mAID-mClover cell line that allows endogenous Ki-67 to be both visualized (via mClover) and targeted for rapid proteasomal degradation (via the mAID degron) was generated [52]. The rapid removal of Ki-67, specifically during mitosis, resulted in chromosomes not only becoming coalesced, but also structurally aberrant or "swollen".…”

The p150N domain of chromatin assembly factor-1 regulates Ki-67 accumulation on the mitotic perichromosomal layer

“…At the PCL, Ki-67’s status as a large, charged protein that possesses “surfactant”-like properties keeps individual mitotic chromosomes dispersed rather than aggregated upon nuclear envelope disassembly, thereby ensuring normal kinetics of anaphase progression (Cuylen et al ., 2016). Similarly, acute depletion experiments show that Ki-67 is important for maintaining mitotic chromosome structure and function (Takagi et al ., 2016). Because p150N regulates interphase Ki-67 localization, we investigated whether p150N also affects Ki-67 localization during mitosis.…”