Project Summary This project is designed to overcome key obstacles in understanding how sodium chloride reabsorption along the distal nephron is regulated by diet and disease. The work will focus on the distal convoluted tubule and the thiazide-sensitive NaCl cotransporter, which play key roles in determining total body potassium balance and in regulating arterial pressure. In fact, we have recently identified this segment as comprising a `potassium switch' that modulates both blood pressure and potassium balance through activation and inhibition of NaCl transport. The thiazide-sensitive NaCl cotransporter is activated by low NaCl intake or low potassium intake. Our overarching hypothesis is that WNK kinases must interact to provide full regulation of thiazide-sensitive NaCl transporter function. We now understand much about dietary K+ transporter regulation, but we don't understand as much about how low dietary NaCl intake regulates that protein. We have a new model suggesting that low NaCl activates the transporter by modulating the carboxyl terminus of a key kinase, WNK4. This region is important because it interacts with other WNKs to form a signaling complex. Here, we will examine its role using a mouse model lacking parts of this domain. We will then determine whether angiotensin II, activated during low NaCl intake, stimulates WNK4 directly, using another novel mouse model. We will also determine whether variations in dietary NaCl intake signal to the distal convoluted tubule from the lumen, by altering calcium delivery to this segment. In a second aim, we will investigate how a second form of WNK, called KS-WNK1, interacts with WNK4 to modulate salt transport. We will test the hypothesis that this protein has dual effects, depending on its location in the cell, acting either to stimulate NaCl transport or to inhibit it. We will use state of the art microscopic techniques and novel mouse models to investigate these questions.