An investigation into the functional role of the RhoGAP protein SH3BP1/ARHGAP43 in Autosomal Dominant Polycystic Kidney Disease

RhoGAP 蛋白 SH3BP1/ARHGAP43 在常染色体显性多囊肾病中的功能作用研究

• 批准号: MR/W015579/1
• 负责人: Joshua Griffiths
• 金额: $ 34.43万
• 依托单位: University of Sheffield
• 依托单位国家: 英国
• 项目类别: Fellowship
• 财政年份: 2022
• 资助国家: 英国
• 起止时间: 2022 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FW015579%2F1
• 关键词: investigation; into; functional; role; RhoGAP

Autosomal Dominant Polycystic Kidney Disease (ADPKD) is a potentially life changing and life-threatening disease that affects up to 1:1000 people. The disease causes fluid filled cysts to grow in the kidneys where they continue to grow to huge sizes through a patient's life. These cysts slowly reduce the function of both kidneys until patients reach end stage kidney failure, most commonly by the ages of 50 to 60. When patients reach end stage kidney failure, they will need dialysis or kidney transplant to survive. ADPKD is caused by a problem in a patient's genetic code which leads to defects in the proteins Polycystin 1 (PC1) or Polycystin 2 (PC2). Parents with one of these genetic problems have a 50:50 chance of passing this down to their children and there is currently no cure. There is only one treatment that can delay developing end stage kidney failure, but it has significant side effects and only slows the disease a small amount. Even though we have known what damaged proteins cause ADPKD for a number of years, how damage to these proteins causes cysts to grow remains a mystery. By looking at the cells lining these these cysts we can see they grow and divide faster but also move slower and are less adherent to each other than normal cells. However, how PC1 or PC2 damage leads to these changes in cells is not currently know.This project will aim to understand what the functions of PC1 and PC2 do when not defective and how defects in these proteins lead to the known changes in cell behaviour and therefore cyst growth. We have newly discovered that PC1 interacts with another protein, SH3BP1. This protein has been shown to be important for cell movement and cell to cell adhesion. Further, it is also important in the uptake into the cells of certain receptors that stimulate cell growth after being activated. This process has only recently been discovered and its potential role in ADPKD has not yet been explored. SH3BP1 provides an exciting potential explanation for why defective PC1 leads to the changes in cells we see under the microscope and therefore how cysts develop.I will use lab-based techniques to establish what parts of PC1 and SH3BP1 interact specifically and if this interaction changes how either protein works. I will also be able to screen for any other proteins that may also be involved in this interaction using mass spectrometry. Using high powered microscopy, I will look at how specific growth receptors are taken up by cells in real time and evaluate if this is different in cells with damaged PC1 or SH3BP1. Finally, to confirm the role of the interaction of PC1 and SH3BP1 in cyst growth, I will see if increasing or decreasing SH3BP1 in cells with damaged PC1 will reduce the growth of cysts in 3D. By completing this project, we will have a better understanding of how cysts grow in ADPKD and this may lead to better treatments for the disease which are desperately needed.

常染色体显性遗传性多囊肾病（ADPKD）是一种潜在的改变生命和威胁生命的疾病，影响高达1：1000的人。这种疾病导致充满液体的囊肿在肾脏中生长，在患者的一生中继续生长到巨大的尺寸。这些囊肿会慢慢降低双肾的功能，直到患者达到终末期肾衰竭，最常见的是50至60岁。当患者达到终末期肾衰竭时，他们将需要透析或肾移植才能生存。ADPKD是由患者遗传密码中的问题引起的，该问题导致多囊蛋白1（PC1）或多囊蛋白2（PC2）的缺陷。患有这些遗传问题的父母有50：50的机会将其遗传给他们的孩子，目前还没有治愈方法。只有一种治疗方法可以延缓终末期肾衰竭的发展，但它有明显的副作用，只能减缓疾病的发展。尽管我们已经知道是什么受损的蛋白质导致ADPKD多年，但这些蛋白质的损伤如何导致囊肿生长仍然是一个谜。通过观察这些囊肿内的细胞，我们可以看到它们生长和分裂得更快，但移动也更慢，并且比正常细胞更少相互粘附。然而，PC1或PC2损伤如何导致细胞中的这些变化目前尚不清楚。该项目旨在了解PC1和PC2在没有缺陷时的功能，以及这些蛋白质的缺陷如何导致细胞行为的已知变化，从而导致囊肿生长。我们新发现PC 1与另一种蛋白SH3BP 1相互作用。这种蛋白质已被证明是重要的细胞运动和细胞与细胞的粘附。此外，它在某些受体被激活后刺激细胞生长的细胞摄取中也很重要。这一过程最近才被发现，其在ADPKD中的潜在作用尚未被探索。SH3BP1提供了一个令人兴奋的潜在解释，为什么有缺陷的PC 1导致我们在显微镜下看到的细胞的变化，因此囊肿如何发展。我将使用基于实验室的技术来确定PC 1和SH3BP1的哪些部分具体相互作用，以及这种相互作用是否改变了蛋白质的工作方式。我也将能够筛选任何其他蛋白质，也可能参与这种相互作用使用质谱。使用高倍显微镜，我将观察特定的生长受体是如何被细胞在真实的时间内吸收的，并评估这在PC 1或SH 3BP 1受损的细胞中是否不同。最后，为了证实PC1和SH3BP1的相互作用在囊肿生长中的作用，我将观察在PC1受损的细胞中增加或减少SH3BP1是否会减少3D中囊肿的生长。通过完成这个项目，我们将更好地了解囊肿如何在ADPKD中生长，这可能会导致更好地治疗迫切需要的疾病。