Shugoshin Prevents Dissociation of Cohesin from Centromeres During Mitosis in Vertebrate Cells (original) (raw)
Figure 3
Sgo1 Depletion Causes Precocious Sister Separation and Mitotic Arrest
Synchronised HeLa cells were transfected with Sgo1 siRNA or dH2O, harvested at 2-h intervals following release and examined by chromosome spreading and Giemsa staining. 100 cells were scored for mitotic index, and 100 mitotic cells were classified into seven categories based on chromosome configuration as exemplified in (B) for each time point.
(A) The frequency of each category of mitotic cell is given as a percentage of total cell numbers, such that the sum of each column represents the mitotic index.
(B) Representative pictures of seven categories of mitotic cells. Chromosome spreads: (a) prophase; (b) metaphase/metaphase-like; (c) anaphase; and (d) telophase. (e) Early phase of precocious sister disjunction. At this stage, sisters are beginning to separate and some or all presumptive sister pairs are still discernible. Arrowheads indicate chromosomes whose centromeric cohesion seems to be lost. (f) Later phases of sister chromatid separation. Sister pairs at this stage are no longer discernible, remnants of the metaphase plate are still visible, and sisters have not yet hypercondensed. (g) Scattered single chromatids. Sisters are completely separated and distributed randomly in relation to one another, individual chromatids are hypercondensed, giving a “curly” appearance. Note that chromatid separation in normal anaphases (c) are different from precocious sister chromatid separation (e–g), in that disjoined and paired chromatids coexist in the same cell.
(C) A portion of the cells harvested for the analysis in Figure 3A and 3B were ethanol-fixed and their DNA content was analysed by flow cytometry.