Defining the functional roles of the enigmatic G protein-coupled receptor GPR35

G蛋白偶联受体（GPCR）是细胞表面蛋白，在允许细胞对外部提示和信号做出反应方面发挥关键作用。它们通常被认为是通过激活一组所谓的G蛋白中的一种或多种来实现这一目的。由于在控制对许多激素和神经递质的反应中的这些关键作用，大量的GPCR是目前使用的药物的分子靶标，并且一般来说，GPCR被认为是“易处理的”，即激活或阻断其激活的分子可以被发现并开发为潜在的新药。然而，有许多GPCR的特征很差，它们在生理功能控制中的作用是未知或不确定的。在这种情况下，它们长期成为新药靶点的可能性，往往部分取决于在编码气相聚合酶链式反应的基因已被消除（敲除）的小鼠中产生的效应，或者在有这种证据的情况下，取决于在人类中编码气相聚合酶链式反应的基因序列变异与患病可能性有关的研究中产生的效应。GPR 35就是这样一种GPCR。已知它存在于结肠中，并且蛋白质序列的变化与下肠道炎性疾病（包括溃疡性结肠炎）的潜在发展有关。GPR 35在炎症调节中也可能具有更普遍的作用。例如，一些能够激活GPR 35的配体已经成为用于治疗气道/肺部炎性疾病哮喘的有用药物。要更好地理解GPR 35是否可以成为新药的有用靶点，有两个主要挑战。首先，虽然有两种化学物质可以阻断人类形式的GPR 35的功能，但它们都不适用于大鼠或小鼠版本的GPR 35。这意味着在啮齿动物生理学和疾病模型中探索GPR 35的功能是非常困难的，尽管这些模型对于支持建立一个可能值得在人类患者中进行测试的案例至关重要。其次，由于GPR 35的信号转导机制既不寻常又缺乏探索，因此不清楚GPR 35产生的各种信号中哪一种可能最适合模拟或阻断以治疗疾病。在拟议的研究中，我们计划使用各种高度创新的方法来克服这些挑战。我们使用了一种称为“基因编辑”的技术来开发一系列细胞系，其中只有GPCR可以产生的信号子集才能被诱导。这些将使我们能够评估GPR 35的不同激活剂是否可能在生理系统中引起不同的影响。第二个关键方法将涉及生产转基因（即基因改造）小鼠。总的来说，小鼠和人类有一组非常相似的基因。然而，在某些情况下，可以激活或阻断人受体的配体对小鼠受体不具有相同的作用。这就是GPR 35的情况。因此，我们计划将小鼠的GPR 35基因替换为人类的相应基因，从而使小鼠“人性化”。这将产生小鼠，其中对GPR 35配体的反应将类似于我们在人类中激活或阻断GPR 35时所预期的反应，并将更清楚地了解GPR 35配体如何影响人类疾病治疗的发展。虽然该申请的主要目标是开发加深我们对GPR 35作用及其功能的理解的方法，但所获得的结果将极大地影响未来关于该受体是否可能成为新药开发新靶点的决定。

G protein-coupled receptors (GPCRs) are cell surface proteins that play key roles in allowing cells to respond to external cues and signals. They are routinely considered to do so by activating one or more of a group of so called G proteins. Because of these key roles in controlling responses to many hormones and neurotransmitters a substantial number of GPCRs are the molecular targets for currently employed medicines and, in general, GPCRs are considered 'tractable' i.e. that molecules that activate or block their activation can be found and developed as potential new medicines. However, there are a number of GPCRs that are poorly characterised and the roles they play in the control of physiological functions are unknown or uncertain. In such cases the potential for them to be targets for new medicines in the longer term is often assessed partially by the effects produced in mice in which the gene encoding the GPCR has been eliminated (knocked-out) or, where such evidence is available, in studies in which variations in the sequence of the gene encoding the GPCR in humans is linked to the potential to develop a disease. GPR35 is such a GPCR. It is known to be present in the colon and variations in sequence of the protein have been linked to the potential development of inflammatory diseases of the lower gut including Ulcerative Colitis. There may also be a more general role for GPR35 in the regulation of inflammation. For example, some ligands that are able to activate GPR35 have been useful medicines for the treatment of airway/lung inflammatory disease asthma. There are two major challenges to better understand if GPR35 could be a useful target for new medicines. Firstly, although there are two chemicals available that can block the function of the human form of GPR35, neither of them work at either the rat or mouse versions of GPR35. This means that it is very difficult to explore the function of GPR35 in rodent models of physiology and disease although such models are vital to provide support to build a case that this might be worth testing in human patients. Secondly, as the mechanisms of signal transduction by GPR35 are both unusual and poorly explored it is unclear which of the various signals generated by GPR35 might be most appropriate to mimic or block to treat disease. In the proposed studies we plan to use a variety of highly innovative approaches to overcome these challenges. We have used a technique called 'gene-editing' to develop a range of cell lines in which only subsets of signals that can be generated by a GPCR are actually induced. These will allow us to assess if different activators of GPR35 are likely to cause different effects in physiological systems. The second key approach will involve the production of transgenic (i.e. genetically modified) mice. Overall, mice and humans have a very similar set of genes. However, in some circumstances ligands that can activate or block a human receptor do not have the same effect at the mouse receptor. This is the case for GPR35. As such, we plan to 'humanise' mice, by replacing the mouse gene for GPR35 with the equivalent gene from humans. This will produce mice in which the responses to GPR35 ligands will be akin to those we would anticipate if we activated or blocked GPR35 in humans and will provide a much clearer picture of how GPR35 ligands might affect the development of treatment of diseases in humans. Although the key objectives of the application are to develop approaches that will deepen our understanding of the roles of GPR35 and how it functions, the results obtained will greatly influence future decisions on whether this receptor might become a new target for the development of novel medicines.