Kif11-haploinsufficient oocytes reveal spatially differential requirements for chromosome biorientation in the spindle

Bipolar spindle assembly and chromosome biorientation are prerequisites for chromosome segregation during cell division. The kinesin motor KIF11 (also widely known as Eg5) drives spindle bipolarization by sliding antiparallel microtubules bidirectionally, elongating a spherical spindle into a bipolar-shaped structure in acentrosomal oocytes. During meiosis I, this process stretches homologous chromosome pairs, establishing chromosome biorientation at the spindle equator. The quantitative requirement for KIF11 in acentrosomal spindle bipolarization and homologous chromosome biorientation remains unclear. Here, using a genetic strategy to modulate KIF11 expression levels, we show that Kif11 haploinsufficiency impairs spindle elongation, leading to the formation of a partially bipolarized spindle during meiosis I in mouse oocytes. While the partially bipolarized spindle allows chromosome stretching in the inner region of its equator, it fails to do so in the outer region, where merotelic kinetochore-microtubule attachments are favored to form. These findings demonstrate the necessity of biallelic functional Kif11 for bipolar spindle assembly in acentrosomal oocytes and reveal a spatially differential requirement for homologous chromosome biorientation within the spindle. Overall design: To investigate the quantitative requirements for KIF11 in mouse oocytes, we used a strategy of heterozygous constitutive deletion and meiotic-specific conditional deletion of the Kif11 gene. For conditional deletion, we crossed a newly established floxed Kif11 allele (Kif11fl) with the meiosis-specific Spo11-Cre and the oocyte-specific Gdf9-Cre and Zp3-Cre recombinases. Offspring derived from mice carrying the conditional deletion allele were used to establish a constitutive deletion allele of Kif11 (Kif11del) for the Spo11-Cre +/- background and Kif11 flox/flox for the Gdf9-Cre or Zp3-Cre +/- background. We then estimated the expression levels of full-length Kif11 mRNA and any residual Kif11 in isolated mature oocytes using poly(A) RNA sequencing followed by counting of reads corresponding to the first and last exon of the Kif11 gene, respectively.