Researchers have developed a method to design drugs that efficiently block rather than activate critical cell receptors which may help avoid adverse medication outcomes.
Department of Medicine
Brown Laboratory: Dennis Brown, PhD
Explore This Research Lab
About the Lab
Our research examines cellular mechanisms and physiological signals responsible for regulating the expression of physiologically important transporting proteins in plasma membranes of kidney epithelial cells. We are especially interested in the intracellular trafficking pathways followed by the vasopressin sensitive water channel, aquaporin 2 (AQP2), in normal function and in water balance disorders such as hereditary and acquired nephrogenic diabetes insipidus (NDI). Principal cells in the collecting duct (CD) respond to vasopressin by increasing the water permeability of their apical plasma membranes. This process involves AQP2 phosphorylation events that control the insertion (exocytosis) and removal (endocytosis) of AQP2 to reversibly shift its location from intracellular vesicles to the plasma membrane. These events are essential for urine concentration to occur. We use both in vivo studies and in vitro transfected epithelial cells in which water channel shuttling has been reconstituted by transfection with AQP2 cDNA. A major current project is to use high throughput screening of chemical libraries to discover new drugs that mimic vasopressin action to correct NDI. Another major interest is acid base transport by CD intercalated cells (IC), which are responsible for distal acid secretion. IC use a similar but distinct vesicle shuttling mechanism to control the number of proton pumps (a vacuolar H+ATPase) in their plasma membrane. This regulates acid secretion and allows IC to help kidneys eliminate an acid load. In a recent study, we performed a protemic analysis to produce a detailed map of the V-ATPase “interactome” and we identified several new regulatory proteins that associate with the V-ATPase to regulate its function. These are now being dissected using convergent cell, molecular and physiological techniques. Our methods include high resolution immunogold electron microscopy and confocal fluorescence imaging; biochemical assays of acidification and water transport in cells and isolated vesicles; enzyme activity assays; molecular biology approaches such as siRNA knockdown; whole animal water and electrolyte balance studies using wild type and genetically engineered mice. While our data are specifically relevant to renal function, the work is also of significance to the biology of epithelial cells in general.
Yui N, Lu HA, Chen Y, Bouley R and Brown, D. Continuous basolateral targeting and microtubule dependent transcytosis of the aquaporin-2 water channel. Am J Physiol Cell Physiol. 2013; 304: C38-C48.
Feinstein TN, Yui N, Webber MJ, Wehbi VL, Stevenson HP, King JD Jr, Hallows KR, Brown D, Bouley R, and Vilardaga J-P. Noncanonical control of vasopressin receptor type 2 signaling by retromer and arrestin. J Biol Chem. 2013; 288: 27849-27860.
Breton S and Brown D. Regulation of luminal acidification by the V-ATPase. Physiology. 2013; 28: 318-329.
Nomura N, Nunes P, Bouley R, Nair AV, Shaw S, Ueda E, Pathomthongtaweechai N, Lu HA and Brown D. High throughput screening identifies AG-490 as a stimulator of aquaporin 2 membrane expression and urine concentration. Am J Physiol Cell Physiol. 2014; 307: C597-C605.
Azroyan A, Cortex-Retamozo V, Bouley R, Ruan YC, Kiselev E, Jacobson KA, Pittet MJ, Brown D, and Breton S. Renal intercalated cells sense and mediate sterile inflammation via the P2Y14 receptor. PLoS ONE. 2015; 10(3); e0121419.
Merkulova M, Paunescu TG, Azroyan A, Marshansky V, Breton S and Brown D. Mapping the H+(V)-ATPase interactome: identification of proteins involved in trafficking, folding, assembly and phosphorylation. Sci Rep. 2015; 5: 14827.
Cheung PW, Nomura N, Nair AV, Pathomthongtaweechai N, Ueberdiek L, Lu HA, Brown D. and Bouley R. EGF receptor inhibition by erlotinib increases AQP2-mediated water reabsorption. J Am Soc Nephrol. 2016, in press.
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About the Nephrology Division
The Division of Nephrology at Massachusetts General Hospital is a leading provider of services for patients with kidney disease, including diagnosis and management of kidney diseases and medical management of renal transplantation.