Interrogating cell division errors using random and chromosome-specific missegregation approaches

Figures & data

Table 1. Summary of available tools and approaches to study cell division defects and/or chromosome segregation errors.

Approach		Inhibited target	Error efficiency	Initial manipulation required?	Inducible?	Prolonged mitosis required?	Trackable system?
Random	Nocodazole	Microtubules	Low	No	Yes	Yes	No
	Colcemid	Microtubules	Low	No	Yes	Yes	No
	Taxol	Microtubules	Low	No	Yes	Yes	No
	Monastral	Eg5	Low	No	Yes	Yes	No
	STLC	Eg5	Low	No	Yes	Yes	No
	GSK923295	CENP-E	Low	No	Yes	Yes	No
	Reversine	Mps1	High	No	Yes	No	No
	AZ3146	Mps1	High	No	Yes	No	No
Selective	Human artificial chromosome	HAC centromere	High	Yes	Yes	No	Yes
	Microcell-mediated chromosome transfer	None	N/A	Yes	No	No	Yes
	CENP-A replacement	Y centromere	High	Yes	Yes	No	Yes

Figure 1. Swapping out CENP-A at the centromere with an inducible gene replacement strategy. Gene editing is first used to add an auxin-inducible degron (AID) sequence to endogenous CENP-A, which undergoes rapid TIR1 E3 ubiquitin ligase-dependent degradation in the presence of the plant hormone auxin. Degraded CENP-A^AID is then rescued by a doxycycline-inducible gene encoding a CENP-A/histone H3 carboxy-tail chimera (CENP-A^C-H3) that does not support kinetochore assembly specifically on the Y centromere, thereby producing Y chromosome-selective segregation errors in the subsequent mitosis.

Figure 2. A catastrophic fate for centromere-inactivated Y chromosomes. Following centromere inactivation using the CENP-A replacement strategy, the Y chromosome fails to properly congress during metaphase and subsequently missegregates in anaphase to produce aneuploid daughter cells with loss, gains, or micronucleation of the Y chromosome. When sequestered in a micronucleus, the Y chromosome undergoes extensive fragmentation in the next mitosis. The resulting fragments incorporate back into the nucleus for subsequent re-ligation by classical non-homologous end joining during interphase.