Coupling of organic anion transport to the glutamate gradient by OATs and OATPs

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The organic anion transporter (OAT) and functionally related organic anion transporting peptide (OATP) families transport a wide range of biologically and pharmacologically important molecules into and out of cells. They transport hormones, metabolites, waste products and many drugs including aspirin, antibiotics, anticancer, cholesterol and blood pressure lowering drugs. They mediate the intestinal absorption of drugs into the body, the transport these drugs into target cells and finally the removal of these drugs from the body via the kidneys and liver. For example, the reason that some medications cannot be taken with grapefruit juice is that it inhibits specific OATPs. This project will investigate how these transporters obtain the energy they require from chemical gradients to undertake their essential biological and pharmacological roles.OAT and OATPs can transport substances both into and out of cells. If you want to pump a drug into a cell, transporting in both directions is inefficient (as what comes in can go straight out again). However, by coupling transporter activity to a second substrate it is possible to mediate transfer of the drug in one direction (as the substance of interest is taken up but cannot get out again because the second substrate is using the transporter). For this to work the second substrate must have a higher concentration within the cell than the substance in question. Glutamate concentrations within the cell are 50-100 x higher than in plasma and we have found that OAT4 and OATP2B1 transport glutamate. Thus the glutamate gradient could drive uptake of drugs and other molecules transported by OAT4 and OATP2B1. In an analogous manner to the release of water stored behind a dam, when the glutamate is released from the cell the potential energy stored in the gradient can be captured and used to drive the uptake of drugs.There are multiple OAT and OATP transporters many of which transport the same molecules. If some of these transporters are coupled to the glutamate gradient, and some are not, then even though they transport the same substrates they will act in different ways and play different roles in the body. Those being coupled to the glutamate gradient (e.g. OAT4) would mediate uptake of drugs and hormones while those which are not would be more likely to mediate efflux. The first aim of this project will be to determine which other members of the OAT and OATP families transport glutamate. The fact that OAT4, OATP2B1 or any other OAT/OATP, transports glutamate does not necessarily mean that its activity will be coupled to the glutamate gradient; this will depend on the kinetics of glutamate transfer, the presence of other substrates and the subcellular localisation of glutamate. The second aim of this project will be to determine when a specific substrate is taken up, then what intracellular molecules is it exchanged for in human cells. It can become difficult to predict how a transporter will function if its activity depends on the concentrations of multiple different substrates on two sides of a biological membrane. It is even more complicated when there are different transporters for the same substrates in the same membrane or in apical and basal membranes of an epithelial barrier. The third aim of this project is to develop a physiologically based mathematical model of how these transporters work. This will allow us to predict how they will function together and how this will affect the transport of their endogenous substrates and of therapeutic agents. The OAT and OATP families play essential biological roles and are important for our understanding of normal cellular function and drug pharmacology. This project explores the extent to which glutamate, which has a large trans-membrane gradient compared to other OAT/OATP substrates, drives the activity of these transporters and provide mathematical models which will help us explain and predict their function throughout the body.