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 #
1R01DK084059-01A1
Application #
7792629
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
2010-04-01
Budget End
2011-03-31
Support Year
1
Fiscal Year
2010
Total Cost
$380,000
Indirect Cost
Name
Washington University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
068552207
City
Saint Louis
State
MO
Country
United States
Zip Code
63130
Milatz, Susanne; Himmerkus, Nina; Wulfmeyer, Vera Christine et al. (2017) Mosaic expression of claudins in thick ascending limbs of Henle results in spatial separation of paracellular Na+ and Mg2+ transport. Proc Natl Acad Sci U S A 114:E219-E227
Gong, Yongfeng; Hou, Jianghui (2017) Claudins in barrier and transport function-the kidney. Pflugers Arch 469:105-113
Gong, Yongfeng; Sunq, Abby; Roth, Robyn A et al. (2017) Inducible Expression of Claudin-1 in Glomerular Podocytes Generates Aberrant Tight Junctions and Proteinuria through Slit Diaphragm Destabilization. J Am Soc Nephrol 28:106-117
Gong, Yongfeng; Himmerkus, Nina; Sunq, Abby et al. (2017) ILDR1 is important for paracellular water transport and urine concentration mechanism. Proc Natl Acad Sci U S A 114:5271-5276
Cain, Matthew D; Salimi, Hamid; Gong, Yongfeng et al. (2017) Virus entry and replication in the brain precedes blood-brain barrier disruption during intranasal alphavirus infection. J Neuroimmunol 308:118-130
Hou, Jianghui; Baker, Lane A; Zhou, Lushan et al. (2016) Viral interactions with the blood-brain barrier: old dog, new tricks. Tissue Barriers 4:e1142492
Penton, David; Czogalla, Jan; Wengi, Agnieszka et al. (2016) Extracellular K+ rapidly controls NaCl cotransporter phosphorylation in the native distal convoluted tubule by Cl- -dependent and independent mechanisms. J Physiol 594:6319-6331
Hou, Jianghui (2016) Claudins and mineral metabolism. Curr Opin Nephrol Hypertens 25:308-13
Hou, Jianghui (2016) Paracellular transport in the collecting duct. Curr Opin Nephrol Hypertens 25:424-8
Gong, Yongfeng; Himmerkus, Nina; Plain, Allein et al. (2015) Epigenetic regulation of microRNAs controlling CLDN14 expression as a mechanism for renal calcium handling. J Am Soc Nephrol 26:663-76

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