Altered microtubule plus end assembly during mitosis as a key trigger for chromosomal instability in human cancer cells

在有丝分裂细胞分裂过程中，整个染色体的永久获得或丢失被称为染色体不稳定性（CIN），是人类癌症的一个主要标志。CIN导致进化的非整倍体，这被认为提供了快速适应能力，从而可能支持肿瘤发生和肿瘤进展。尽管这种高度流行的癌症表型很重要，但CIN的分子机制仍然难以捉摸。我们最近的工作现在揭示了人类癌细胞中CIN的关键和广泛的触发因素。事实上，有丝分裂纺锤体中微管加端动力学的增加可以触发短暂的有丝分裂纺锤体异常，导致微管-着丝点错误附着和染色体错误分离。我们发现致癌的Aurora-A激酶是该表型的关键诱导因子，肿瘤抑制基因CHK2和BRCA1是Aurora-A的负调节因子。然而，目前尚不清楚CHK2-BRCA1肿瘤抑制通路如何抑制Aurora-A的活性，以及有丝分裂中心体上Aurora-A激酶的高活性如何导致微管+端组装增加和染色体错分离。考虑到增加的微管加端组装是人类癌细胞染色体不稳定性的基础，了解有丝分裂过程中确保适当的微管动力学的分子机制以及揭示微管加端组装的增加如何驱动染色体错分离和CIN是至关重要的。这些问题将是我们研究计划的重点。特别是，我们将解决CHK2-BRCA1肿瘤抑制途径如何抑制有丝分裂中心体上的Aurora-A活性的问题，我们将分析中心体Aurora-A活性的增加是如何引起纺锤体组装和定位缺陷的，这些缺陷随后可能促进超稳定和错误的微管-着丝点附着体的产生。此外，我们的目标是系统地识别可能在人类癌症中改变的微管加端组装的新调节因子，从而可能代表癌症中的新CIN基因。为此，我们将使用基于我们最近在实验室开发的一种新的表型分析的大规模siRNA筛选来强有力地识别微管组装因子。新发现的介导微管组装率增加和染色体不稳定性的基因将对其在有丝分裂过程中的作用及其在人类癌症中的改变进行详细的表征。因此，我们的研究计划旨在在分子水平上表征微管加端组装增加的表型作为CIN的关键机制，并确定在人类癌细胞中导致这种有趣表型的基因。这些CIN基因可能是抑制肿瘤促进CIN表型的抗癌治疗的有吸引力的靶点。

The perpetual gain or loss of whole chromosomes during mitotic cell division is known as chromosomal instability (CIN) and represents a major hallmark of human cancer. CIN results in evolving aneuploidy, which is thought to provide rapid adaptation capabilities and thereby might support tumorigenesis and tumor progression. Despite the importance of this highly prevalent cancer phenotype the molecular mechanisms underlying CIN remained elusive. Our most recent work now revealed a key and wide-spread trigger for CIN in human cancer cells. In fact, increased microtubule plus end dynamics within mitotic spindles can trigger transient mitotic spindle abnormalities leading to erroneous microtubule-kinetochore attachments and chromosome missegregation. We identified the oncogenic Aurora-A kinase as a key inducer of this phenotype and the tumor suppressor genes CHK2 and BRCA1 as negative regulators of Aurora-A. However, it is not clear how the CHK2-BRCA1 tumor suppressor pathway can restrain the activity of Aurora-A and how the hyper-activity of the Aurora-A kinase at mitotic centrosomes can lead to increased microtubule plus end assembly and chromosome missegregation. Given the fact that increased microtubule plus end assembly underlies chromosomal instability in human cancer cells it is pivotal to understand the molecular mechanisms ensuring proper microtubule dynamics during mitosis and to reveal how an increase in microtubule plus end assembly can drive chromosome missegregation and CIN. These questions will be in focus of our research proposal. In particular, we will address the question how the CHK2-BRCA1 tumor suppressor pathway can restrain Aurora-A activity at mitotic centrosomes and and we will analyze how an increase in centrosomal Aurora-A activity is causing spindle assembly and positioning defects that subsequently might facilitate the generation of hyper-stable and erroneous microtubule-kinetochore attachments. Moreover, we aim to systematically identify novel regulators of microtubule plus end assembly that might be altered in human cancer and thus, might represent novel CIN genes in cancer. For this, we will use large-scale siRNA screens based on a new phenotypic assay that we recently developed in our lab to robustly identify microtubule assembly factors. Newly identified genes that mediate an increase in microtubule assembly rates and chromosomal instability will be subject to detailed characterization for their role during mitosis and for their alteration in human cancer. Thus, our research plan aims to characterize the phenotype of increased microtubule plus end assembly as a key mechanism for CIN at a molecular level and to identify genes that contribute to this intriguing phenotype in human cancer cells. Those CIN genes might represent attractive targets for anti-cancer therapy to suppress the tumor promoting CIN phenotype.