(provided by candidate): Epithelial sodium channels (ENaC) play a pivotal role in salt and water homeostasis which is important for maintaining normal blood pressure in humans. ENaC is a heteromeric protein consisting of three subunits alpha, beta and gamma. The major systemic hormone modulating ENaC activity is the steroid hormone aldosterone. Although, understanding aldosterone's action would help understand the control of blood pressure by aldosterone, the mechanism of aldosterone control of ENaC activity is unclear. The overall aim of this study is to elucidate the mechanisms by which aldosterone stimulates ENaC. The increase in ENaC message or protein levels in response to aldosterone are not consistent across different species or tissues. In general, there is no good evidence for an increase in ENaC message or translation rate being related to the increase in transport. Nonetheless, electro-physiological data suggests that an increase in the number of functional channels at the cell surface contributes to aldosterone's action. Since new protein synthesis does not account for the increase in channel number, we propose that aldosterone increases the rate of ENaC assembly and insertion from a pre-existing intracellular pool into the apical membrane, or alternately decreases the rate of channel retrieval and degradation from the apical membrane, Our initial data support both of these hypothesis, Therefore, we will use both biochemical and molecular biology tools to study the effects of aldosterone on ENaC trafficking and degradation. We will determine the effect of aldosterone in a renal cell line, A6, expressing native ENaC (one of the few systems in which ENaC trafficking can be studied in native epithelial cells) and in transfected Chinese Hamster Ovary (CHO) cells. Transfected CHO cells are a useful system because transfected ENaC subunits lack steroid response elements; and thus, there can be no direct ENaC induction expression by aldosterone. To further, characterize aldosterone effects on ENaC degradation, we will determine the role of the hormone in the regulation of various proteins involved in the ENaC protein degradative pathway. One major protein in ENaC proteolytic pathway is Nedd4. We will study the effect of aldosterone on Nedd4 activity and also on Nedd4 message and protein. This work should provide a better understanding of the activators of aldosterone and possibly provide new therapeutic targets for the reduction of hypertension.
|Malik, B; Yue, Q; Yue, G et al. (2005) Role of Nedd4-2 and polyubiquitination in epithelial sodium channel degradation in untransfected renal A6 cells expressing endogenous ENaC subunits. Am J Physiol Renal Physiol 289:F107-16|