Phosphoproteomic analysis reveals regulatory mechanisms at the kidney filtration barrier.

Abstract

Diseases of the kidney filtration barrier are a leading cause of ESRD. Most disorders affect the podocytes, polarized cells with a limited capacity for self-renewal that require tightly controlled signaling to maintain their integrity, viability, and function. Here, we provide an atlas of in vivo phosphorylated, glomerulus-expressed proteins, including podocyte-specific gene products, identified in an unbiased tandem mass spectrometry-based approach. We discovered 2449 phosphorylated proteins corresponding to 4079 identified high-confidence phosphorylated residues and performed a systematic bioinformatics analysis of this dataset. We discovered 146 phosphorylation sites on proteins abundantly expressed in podocytes. The prohibitin homology domain of the slit diaphragm protein podocin contained one such site, threonine 234 (T234), located within a phosphorylation motif that is mutated in human genetic forms of proteinuria. The T234 site resides at the interface of podocin dimers. Free energy calculation through molecular dynamic simulations revealed a role for T234 in regulating podocin dimerization. We show that phosphorylation critically regulates formation of high molecular weight complexes and that this may represent a general principle for the assembly of proteins containing prohibitin homology domains.

• Authors: Markus M Rinschen, Xiongwu Wu, Tim König, Trairak Pisitkun, Henning Hagmann, Caroline Pahmeyer, Tobias Lamkemeyer, Priyanka Kohli, Nicole Schnell, Bernhard Schermer, Stuart Dryer, Bernard R Brooks, Pedro Beltrao, Marcus Krueger, Paul T Brinkkoetter, Thomas Benzing
• Journal: Journal of the American Society of Nephrology : JASN (J Am Soc Nephrol) Vol. 25 Issue 7 Pg. 1509-22 (Jul 2014) ISSN: 1533-3450 [Electronic] United States
• PMID: 24511133 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: Copyright © 2014 by the American Society of Nephrology.

Chemical References

• Intracellular Signaling Peptides and Proteins
• Membrane Proteins
• NPHS2 protein
• Phosphoproteins

Topics

• Animals
• Female
• Glomerular Filtration Barrier (physiology)
• Intracellular Signaling Peptides and Proteins (physiology)
• Membrane Proteins (physiology)
• Mice
• Phosphoproteins (analysis, physiology)
• Phosphorylation
• Podocytes (physiology)
• Proteomics