[Research Grant] Juxtamembrane control of DDR1 kinase activity
Ente: Biotechnology and Biological Sciences Research Council
Scadenza: 2021-07-15
Paese: GB
Descrizione
The cells in our body are not passive and static building blocks like bricks in a wall; rather, they constantly monitor and react to their environment. They do this by sending and receiving messages in the form of signalling molecules. In order to perceive a particular signal, a cell has to have appropriate sensors. For messages that are received from the cell's environment, the sensor is often a specialised protein molecule called a receptor tyrosine kinase (RTK). One part of the RTK sticks out from the cell and another part is inside the cell. When the outside part interacts with a signalling molecule, the receptor changes its shape. This causes the inside part to become active and carry out a chemical reaction (phosphorylation) that ultimately changes the cell's behaviour. RTKs control many important functions, such as cell division, and their activity must be tightly controlled in order to prevent the development of diseases, such as cancer. Research into how RTKs are controlled is important for understanding normal human physiology, as well as for understanding what goes wrong in disease. RTKs are the targets of many drugs used in cancer therapy, and basic research of RTKs is required for designing more effective drugs.
This project will establish how the activity of an RTK called DDR1 is controlled. DDR1 instructs cells to change their behaviour when collagen is present. We discovered that a part of DDR1, which we named JM4, is needed for DDR1 signalling activity. When JM4 is missing, DDR1 can still bind to collagen, but no phosphorylation reaction results inside the cell. JM4 is also needed when the phosphorylation reaction is done in a test tube rather than in a cell. Therefore, we believe that JM4 is an important control region that regulates the part of DDR1 that carries out the phosphorylation reaction, which is called the kinase.
In this project, we aim to obtain a detailed understanding of how JM4 controls the kinase activity of DDR1. We will determine which part of the phosphorylation reaction is enhanced by JM4 and whether JM4 pushes the kinase into an active shape. In addition to directly affecting the shape of the kinase, JM4 could also affect its activity indirectly by interacting with another cellular component called Src. We already determined that Src increases DDR1 phosphorylation in cells. In this project, we will determine how Src increases DDR1 kinase activity and whether this occurs with the help of JM4.
This research is important because it may provide the basis for designing novel drugs against faulty DDR1 signalling in human disease. Most drugs against RTKs are designed to block the kinase activity by directly blocking the active part of the kinase. Because cells contain hundreds of other kinases with active parts of similar shape, these types of drugs often lead to serious side effects. JM4 is only found in DDR1 (and a similar protein called DDR2). Understanding the precise role of JM4 in controlling DDR1 kinase a
Receptor tyrosine kinases (RTKs) are an important class of signalling receptors whose dysregulation is associated with disease. The collagen receptor DDR1 is a drug target for kidney disease, fibrosis and many cancers, but in contrast to other RTKs little is known about how the enzymatic activity of DDR1 is regulated. We identified a 26-amino acid segment (JM4) within the DDR1 juxtamembrane region, which positively regulates DDR1 kinase catalytic activity. In this project, we will carry out detailed mechanistic studies in order to define how JM4 controls DDR1 activation.
Using in vitro kinase assays of both isolated cytosolic constructs and full-length DDR1 expressed in cells, we will determine the effect of JM4 on the kinetic parameters of DDR1. Using biophysical techniques, we will investigate the effect of JM4 on DDR1 kinase dimerisation and its affinity for ATP. Using X-ray crystallography, we will determine the structure of the DDR1 kinase with JM4. Furthermore, we will investigate the role of Src in promoting DDR1 kinase activity, characterise the physical interaction of Src with DDR1, and determine whether JM4 provides a binding site for Src. Finally, we will carry out cell biological experiments to define the roles of different Src family kinases in DDR1 activation and downstream signalling.
Collectively, these studies will define the mechanism by which JM4 controls DDR1 kinase catalytic activity. This information will help in the design of more selective drugs agai
Settori: National Heart and Lung Institute
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