We have identified a new type of growth factor called Ngal (lipocalin-2) which is a member of the lipocalin- superfamily. It is known that lipocalins bind and transport small chemicals ligands, but little has beenknown about Ngal, and some of its presumptive ligands have been disproved. We purified Ngal from liters of media conditioned by an embryonic cell line, which revealed a growthfactor with a reddish color. The cloned protein was also colored and x-ray crystallography of the cloned protein (R. Strong, Fred Hutchinson Cancer Research Center) showed that it contained a siderophore:Fe complex that it had bound during cloning. Using recombinant Ngal we could induce growth and conversion of mesenchymeinto epithelia, tubule and nephrons, but only when rNgal was loaded with this siderophore and iron. This suggested that Ngal's inductive action must have been due to its ligand, and we speculate that perhaps a mammalian versionof the siderophore is present in vivo. Ngal is one of the most overexpressed proteins in ischemic disease of the kidney (>1000 fold increases). It is also the earliest known protein to be increased in the urine and in the serum of ischemic and toxicdiseases of the kidney. In situ hybridization shows that Ngal is massively expressed by TALH and collecting ducts. In this grant we propose to study the activity of the endogenous Ngal protein. We will do this by completing an isolation of a small organic molecule that binds Ngal and binds iron. The Ngal ligand is found in mammalian urine. We will identify the structure of this compound using IR, NMR, Mass spectroscopyand x- ray crystallography. To further examine the function of Ngal in the kidney we have deleted its gene in a tissue specific manner. Because Ngal previously had a protective function in an ischemic model of the kidney, we will examine whether the Ngal knockout is more sensitive to ischemia reperfusion injury. We will test the hypothesis that the Ngal:urine siderophore complex removescatalytic iron and converselythe Ngal knockout contains more catalytic iron and iron mediated cell damage. The knockout will then be rescuedby exogenous Ngal or iron chelators. This work is important because it characterizes a novel growth factorthat is massively expressed by damaged renal epithelia and it evaluates a novel iron transport pathwaythat appears critical to Ngal function.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK073462-05
Application #
8019032
Study Section
Cellular and Molecular Biology of the Kidney Study Section (CMBK)
Program Officer
Hoshizaki, Deborah K
Project Start
2007-01-01
Project End
2011-11-30
Budget Start
2010-12-01
Budget End
2011-11-30
Support Year
5
Fiscal Year
2011
Total Cost
$317,012
Indirect Cost
Name
Columbia University (N.Y.)
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
621889815
City
New York
State
NY
Country
United States
Zip Code
10032
Barasch, Jonathan; Hollmen, Maria; Deng, Rong et al. (2016) Disposal of iron by a mutant form of lipocalin 2. Nat Commun 7:12973
Bao, Guan-Hu; Barasch, Jonathan; Xu, Jie et al. (2015) Purification and Structural Characterization of "Simple Catechol", the NGAL-Siderocalin Siderophore in Human Urine. RSC Adv 5:28527-28535
Allred, Benjamin E; Rupert, Peter B; Gauny, Stacey S et al. (2015) Siderocalin-mediated recognition, sensitization, and cellular uptake of actinides. Proc Natl Acad Sci U S A 112:10342-7
Bao, Guan-Hu; Ho, Chi-Tang; Barasch, Jonathan (2015) The Ligands of Neutrophil Gelatinase-Associated Lipocalin. RSC Adv 5:104363-104374
Bao, Guan-Hu; Xu, Jie; Hu, Feng-Lin et al. (2013) EGCG inhibit chemical reactivity of iron through forming an Ngal-EGCG-iron complex. Biometals 26:1041-50
Allred, Benjamin E; Correnti, Colin; Clifton, Matthew C et al. (2013) Siderocalin outwits the coordination chemistry of vibriobactin, a siderophore of Vibrio cholerae. ACS Chem Biol 8:1882-7
Singer, E; Markó, L; Paragas, N et al. (2013) Neutrophil gelatinase-associated lipocalin: pathophysiology and clinical applications. Acta Physiol (Oxf) 207:663-72
Correnti, Colin; Richardson, Vera; Sia, Allyson K et al. (2012) Siderocalin/Lcn2/NGAL/24p3 does not drive apoptosis through gentisic acid mediated iron withdrawal in hematopoietic cell lines. PLoS One 7:e43696
Nickolas, Thomas L; Forster, Catherine S; Sise, Meghan E et al. (2012) NGAL (Lcn2) monomer is associated with tubulointerstitial damage in chronic kidney disease. Kidney Int 82:718-22
Sise, Meghan E; Parravicini, Elvira; Barasch, Jonathan (2012) Urinary neutrophil gelatinase associated lipocalin identifies neonates with high probability of sepsis. Pediatrics 130:e1053-4; author reply e1055-7

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