The physiology of paracellular permeation of ions and solutes in the kidney is pivotally important but poorly understood. Claudins are the key components of the paracellular pathway. Defects in claudin function result in a broad range of renal diseases, including hypomagnesemia, hypercalciuria, hypochloremia and salt- sensitive hypertension. Human mutations in claudin-16 and claudin-19 are responsible for the hereditary renal disease FHHNC (Familial Hypomagnesemia with Hypercalciuria and Nephrocalcinosis). We have found that the two claudins co-localize in the thick ascending limb (TAL), interact and form a cation-selective protein complex. We have developed claudin-16 knockdown (KD) and claudin-19 KD mouse lines. Claudin-16 KD animals show chronic renal wasting of magnesium and calcium, developing renal nephrocalcinosis comparable to that seen in human patients with FHHNC. This proposal will study the molecular interaction between claudin-16 and claudin-19, the size and charge selectivity of claudin-16 and claudin-19 channel pore and their collective effects on renal handling of Mg++.
Specific Aim 1 intends to understand how claudin-16 and claudin-19 co-oligomerize and assemble into the tight junction strands. These studies will determine the stoichiometry of claudin-16/claudin- 19 oligomeric complex and identify critical intracellular step in claudin oligomerization using metabolic chase analyses.
Specific Aim 2 intends to elucidate the biophysical properties of the claudin-16 and claudin-19 channels. These studies will measure the paracellular flux of a continuous series of PEG oligomers (of radius 2.8 - 7 ?) to unveil the size selectivity of claudin-16 and claudin-19 channels. These studies will identify key loci of amino acids in claudin-16 and claudin-19 required for their charge selectivity. These studies will test two models of claudin-16 and claudin-19 channel structure and function: (1) claudin-16 and claudin-19 form two parallel homomeric channels each with its own physiologic signature;(2) claudin-16 and claudin-19 form a heteromeric channel with novel properties that require their synergy.
Specific Aim 3 involves careful phenotypic analyses of claudin-16 KD mice, claudin-19 KD (KO) mice and claudin-16 KD + claudin-19 KD (KO) mice. These studies will analyze the renal clearance and transport functions in these mice, and record the electrophysiological properties of claudin-16 and claudin-19 channels in the TAL of these mice using single tubule perfusion techniques. These studies intend to test if the interaction between claudin-16 and claudin-19 is required for normal function of the TAL in vivo.

Public Health Relevance

Kidneys function by initially excreting many salts and small molecules found in the blood, then selectively taking back those that need to be conserved while allowing others to be excreted in the urine. This grant will study claudin functions of cell-cell junctions that provide one of the key pathways (the paracellular pathway) used by the kidney to move salt between urine and blood. Defects in claudin function result in a broad range of renal diseases, including hypomagnesemia, hypercalciuria, hypochloremia and salt- sensitive hypertension.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
3R01DK084059-03S1
Application #
8587125
Study Section
Cellular and Molecular Biology of the Kidney Study Section (CMBK)
Program Officer
Ketchum, Christian J
Project Start
2010-04-01
Project End
2015-03-31
Budget Start
2012-11-30
Budget End
2013-03-31
Support Year
3
Fiscal Year
2013
Total Cost
$1,500
Indirect Cost
$513
Name
Washington University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
068552207
City
Saint Louis
State
MO
Country
United States
Zip Code
63130
Gong, Yongfeng; Hou, Jianghui (2014) Claudin-14 underlies Caýýýýýý-sensing receptor-mediated Caýýýýýý metabolism via NFAT-microRNA-based mechanisms. J Am Soc Nephrol 25:745-60
Gong, Yongfeng; Yu, Miao; Yang, Jing et al. (2014) The Cap1-claudin-4 regulatory pathway is important for renal chloride reabsorption and blood pressure regulation. Proc Natl Acad Sci U S A 111:E3766-74
Hou, Jianghui (2014) The kidney tight junction (Review). Int J Mol Med 34:1451-7
Hou, Jianghui (2013) The role of claudin in hypercalciuric nephrolithiasis. Curr Urol Rep 14:5-12
Hou, Jianghui; Rajagopal, Madhumitha; Yu, Alan S L (2013) Claudins and the kidney. Annu Rev Physiol 75:479-501
Chen, Chiao-Chen; Zhou, Yi; Morris, Celeste A et al. (2013) Scanning ion conductance microscopy measurement of paracellular channel conductance in tight junctions. Anal Chem 85:3621-8
Gong, Yongfeng; Renigunta, Vijayaram; Himmerkus, Nina et al. (2012) Claudin-14 regulates renal Caýýýýýý transport in response to CaSR signalling via a novel microRNA pathway. EMBO J 31:1999-2012
Hou, Jianghui; Goodenough, Daniel A (2010) Claudin-16 and claudin-19 function in the thick ascending limb. Curr Opin Nephrol Hypertens 19:483-8
Hou, Jianghui; Renigunta, Aparna; Yang, Jing et al. (2010) Claudin-4 forms paracellular chloride channel in the kidney and requires claudin-8 for tight junction localization. Proc Natl Acad Sci U S A 107:18010-5