Centrobin regulates the assembly of functional mitotic spindles

Abstract

The proper function of the spindle is crucial to the high fidelity of chromosome segregation and is indispensable for tumor suppression in humans. Centrobin is a recently identified centrosomal protein that has a role in stabilizing the microtubule structure. Here we functionally characterize the defects in centrosome integrity and spindle assembly in Centrobin-depleted cells. Centrobin-depleted cells show a range of spindle abnormalities including unfocused poles that are not associated with centrosomes, S-shaped spindles and mini spindles. These cells undergo mitotic arrest and subsequently often die by apoptosis, as determined by live cell imaging. Co-depletion of Mad2 relieves the mitotic arrest, indicating that cells arrest due to a failure to silence the spindle checkpoint in metaphase. Consistent with this, Centrobin-depleted metaphase cells stained positive for BubR1 and BubR1 S676. Staining with a panel of centrosome markers showed a loss of centrosome anchoring to the mitotic spindle. Furthermore, these cells show less cold-stable microtubules and a shorter distance between kinetochore pairs. These results show a requirement of Centrobin in maintaining centrosome integrity, which in turn promotes anchoring of mitotic spindle to the centrosomes. Furthermore, this anchoring is required for the stability of microtubulekinetochore attachments and biogenesis of tension-ridden and properly functioning mitotic spindle.

Centrobin-mediated Regulation of the Centrosomal Protein 4.1-associated Protein (CPAP) Level Limits Centriole Length during Elongation Stage

“…Our results show that abnormal levels of centrobin are associated with loss of centrosome integrity, defective mitotic spindles with abnormal spindle poles and orientation. In fact this is in agreement with previous reports that showed defective mitotic spindle formation and partial loss of centrosome integrity upon centrobin depletion of centrobin or CPAP (35,51) as well as upon overexpression of CPAP (35). The normal functioning of mitotic spindles is critical for the fidelity of chromosome segregation and tumor suppression.…”

“…Perturbing the centrosome integrity due to short centrioles or abnormal sized centrioles has been shown to affect the bipolar spindle assembly process (35,51), which can contribute to chromosome instability (15,58). Hence, we evaluated the physiological effects of CPAP loss as a consequence of centrobin depletion as well as excess CPAP due to overexpression of centrobin in a mitotic bipolar spindle assembly assay.…”

The RAD51 paralogs ensure cellular protection against mitotic defects and aneuploidy

“…Centrobin is a centriole-associated protein that is required for centriole duplication (Zou et al, 2005). It was recently shown that centrobin-depleted cells have compromised centrosome and spindle integrity, leading to persistent SAC activation (Jeffery et al, 2010). Since RAD51C-deficient hamster cells have an increased number of centrosomes in mitotic cells (Renglin Lindh et al, 2007), we verified whether centrosomes abnormalities might account for persistent SAC activation in siXRCC3 cells by pericentrin staining.…”

Spontaneous slow replication fork progression elicits mitosis alterations in homologous recombination-deficient mammalian cells

“…2A) and centrobin ( Fig. S4), the latter of which is a centriole-associated protein that is required for centriole elongation and stability and regulates functional mitotic spindles (36)(37)(38). We first confirmed with both antibodies that V-C8 and V79SM24 (HR − ) mutants displayed a spontaneous increase in the frequency of mitotic cells bearing MECs compared with untreated HR − proficient cells.…”