Mitotic exit network and human cancer

描述(申请人提供)：癌症遗传不稳定的主要原因之一是有丝分裂错误导致子代细胞之间遗传物质的不平等分离。许多有丝分裂事件以及监控这些事件的检查点在分子水平上尚不清楚。这对于发生在有丝分裂后期到末期转变的事件尤其正确，当细胞解聚它们的姐妹染色单体并重组它们的核时。在酿酒酵母中，调节这些事件的途径被称为有丝分裂退出网络(MEN)。在人类细胞中，人类途径还没有被描述出来。然而，人类蛋白激酶LATS 1与酵母蛋白激酶有很高的序列相似性。LATS 1与有丝分裂有关，但其确切功能尚不清楚。然而，Lats1显然与癌症的发展有关；在苍蝇、小鼠和人类中，LATS 1是一种公认的肿瘤抑制蛋白。我们假设LATS 1在人类细胞中的一条男性途径中发挥作用，该途径的解除调控会导致遗传不稳定(通过杂合性丧失分析来衡量)。将通过执行以下具体目标来解决这一假设： 1.确定LATS 1是酿酒酵母Dbf2蛋白的人类同源基因。我们将研究Lats 1与Dbf2是否具有受Mob蛋白调控的能力。我们将使用siRNA方法和显性负突变进一步研究LATS 1是否在人类细胞的MEN途径中发挥作用。 2.确定LATS 1受hDmal调控。在S.pombe中，男性的激酶由Dmal调节，Dmal是一种假定的泛素连接酶。我们将检查人类LATS 1是否受hDmal调控，hDmal是一种泛素连接酶，与酵母Dmal具有很高的序列相似性。 3.证实LATS 1和hDmal的解除调控增加了杂合性丢失(LOH)的频率。LOH显然与癌症的发生和发展有关。因此，这些研究将有助于确定LATS 1抑癌功能的分子基础。

DESCRIPTION (provided by applicant): One of the major causes of genetic instability in cancer is errors in mitosis leading to unequal segregation of the genetic material between daughter cells. Many of the mitotic events and the checkpoints that monitor these events are yet to be understood at the molecular level. This is particularly true for the events that occur in late mitosis at the anaphase to telophase transition, when cells decondense their sister chromatids and reassemble their nuclei. In S. cerevisiae, the pathway that mediates these events is referred to as the mitotic exit network (MEN). In human cells a MEN pathway has not been characterized. However, a human protein kinase, Lats 1, has high sequence similarity to a yeast MEN kinase. Lats 1 has been implicated in mitosis, but its precise function is unclear. Nevertheless, Latsl is clearly linked to cancer development; in flies, mice and humans, Lats 1 is a well-established tumor suppressor protein. We hypothesize that Lats 1 functions in a MEN pathway in human cells and that deregulation of this pathway leads to genetic instability (as measured by loss-of-heterozygosity analysis). This hypothesis will be addressed by performing the following Specific Aims: 1. Establish that Lats 1 is the human ortholog of the S. cerevisiae Dbf2 MEN kinase. We will examine if Lats 1 shares with Dbf2 the ability to be regulated by Mob proteins. We will further examine whether Lats 1 functions in the MEN pathway in human cells using an siRNA approach and dominant negative mutants. 2. Establish that Lats 1 is regulated by hDmal. In S. pombe the MEN kinases are regulated by Dmal, a putative ubiquitin ligase. We will examine if human Lats 1 is regulated by hDmal, a ubiquitin ligase that has high sequence similarity with yeast Dmal. 3. Establish that deregulation of LATS 1 and hDmal increases the frequency of loss-of-heterozygosity (LOH). LOH is clearly involved in cancer development and progression. Thus, these studies will help identify the molecular basis for the tumor suppressor function of Lats 1.