Haren Laurence, Gnadt Nicole, Wright Michel, Merdes Andreas
Wellcome Trust Centre for Cell Biology, University of Edinburgh, King's Buildings, Edinburgh EH9 3JR, UK.
BMC Res Notes. 2009 Apr 28;2:64. doi: 10.1186/1756-0500-2-64.
NuMA is a protein that has been previously shown to play a role in focusing microtubules at the mitotic spindle poles. However, most previous work relies on experimental methods that might cause dominant side effects on spindle formation, such as microinjection of antibodies, overexpression of mutant protein, or immunodepletion of NuMA-containing protein complexes.
To circumvent these technical problems, we performed siRNA experiments in which we depleted the majority of NuMA in human cultured cells. Depleted mitotic cells show a prolonged duration of prometaphase, with spindle pole defects and with unattached, unaligned chromosomes.
Our data confirm that NuMA is important for spindle pole formation, and for cohesion of centrosome-derived microtubules with the bulk of spindle microtubules. Our findings of NuMA-dependent defects in chromosome alignment suggest that NuMA is involved in stabilizing kinetochore fibres.
核有丝分裂器蛋白(NuMA)是一种先前已被证明在有丝分裂纺锤体极聚焦微管过程中发挥作用的蛋白质。然而,大多数先前的研究依赖于可能对纺锤体形成产生显性副作用的实验方法,如抗体显微注射、突变蛋白过表达或含NuMA蛋白复合物的免疫耗竭。
为了规避这些技术问题,我们在人类培养细胞中进行了小干扰RNA(siRNA)实验,其中我们耗尽了大部分NuMA。耗尽NuMA的有丝分裂细胞显示前中期持续时间延长,伴有纺锤体极缺陷以及未附着、未排列的染色体。
我们的数据证实,NuMA对于纺锤体极的形成以及中心体衍生微管与大部分纺锤体微管的结合很重要。我们发现NuMA依赖的染色体排列缺陷表明,NuMA参与稳定动粒纤维。