Universität zu Köln

Schurek, Eva-Maria, Voelker, Linus A., Tax, Judit, Lamkemeyer, Tobias, Rinschen, Markus M., Ungrue, Denise, Kratz, John E., III, Sirianant, Lalida, Kunzelmann, Karl, Chalfie, Martin, Schermer, Bernhard ORCID: 0000-0002-5194-9000, Benzing, Thomas and Hoehne, Martin (2014). A Disease-causing Mutation Illuminates the Protein Membrane Topology of the Kidney-expressed Prohibitin Homology ( PHB) Domain Protein Podocin. J. Biol. Chem., 289 (16). S. 11262 - 11272. ROCKVILLE: AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC. ISSN 1083-351X

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Abstract

Background: Mutations in the stomatin family protein podocin are the most common genetic cause of proteinuria. Results: A conserved proline residue of podocin is essential for its membrane topology. Conclusion: This study confirms a hairpin-like structure of the membrane-attached PHB domain protein and its significance for cholesterol recruitment. Significance: Podocin(P118L) elucidates the pathogenic implication in kidney disease and identifies a novel family of PHB domain proteins. Mutations in the NPHS2 gene are a major cause of steroid-resistant nephrotic syndrome, a severe human kidney disorder. The NPHS2 gene product podocin is a key component of the slit diaphragm cell junction at the kidney filtration barrier and part of a multiprotein-lipid supercomplex. A similar complex with the podocin ortholog MEC-2 is required for touch sensation in Caenorhabditis elegans. Although podocin and MEC-2 are membrane-associated proteins with a predicted hairpin-like structure and amino and carboxyl termini facing the cytoplasm, this membrane topology has not been convincingly confirmed. One particular mutation that causes kidney disease in humans (podocin(P118L)) has also been identified in C. elegans in genetic screens for touch insensitivity (MEC-2(P134S)). Here we show that both mutant proteins, in contrast to the wild-type variants, are N-glycosylated because of the fact that the mutant C termini project extracellularly. Podocin(P118L) and MEC-2(P134S) did not fractionate in detergent-resistant membrane domains. Moreover, mutant podocin failed to activate the ion channel TRPC6, which is part of the multiprotein-lipid supercomplex, indicative of the fact that cholesterol recruitment to the ion channels, an intrinsic function of both proteins, requires C termini facing the cytoplasmic leaflet of the plasma membrane. Taken together, this study demonstrates that the carboxyl terminus of podocin/MEC-2 has to be placed at the inner leaflet of the plasma membrane to mediate cholesterol binding and contribute to ion channel activity, a prerequisite for mechanosensation and the integrity of the kidney filtration barrier.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Schurek, Eva-Maria UNSPECIFIED UNSPECIFIED UNSPECIFIED Voelker, Linus A. UNSPECIFIED UNSPECIFIED UNSPECIFIED Tax, Judit UNSPECIFIED UNSPECIFIED UNSPECIFIED Lamkemeyer, Tobias UNSPECIFIED UNSPECIFIED UNSPECIFIED Rinschen, Markus M. UNSPECIFIED UNSPECIFIED UNSPECIFIED Ungrue, Denise UNSPECIFIED UNSPECIFIED UNSPECIFIED Kratz, John E., III UNSPECIFIED UNSPECIFIED UNSPECIFIED Sirianant, Lalida UNSPECIFIED UNSPECIFIED UNSPECIFIED Kunzelmann, Karl UNSPECIFIED UNSPECIFIED UNSPECIFIED Chalfie, Martin UNSPECIFIED UNSPECIFIED UNSPECIFIED Schermer, Bernhard UNSPECIFIED orcid.org/0000-0002-5194-9000 UNSPECIFIED Benzing, Thomas UNSPECIFIED UNSPECIFIED UNSPECIFIED Hoehne, Martin UNSPECIFIED UNSPECIFIED UNSPECIFIED
URN:	urn:nbn:de:hbz:38-440606
DOI:	10.1074/jbc.M113.521773
Journal or Publication Title:	J. Biol. Chem.
Volume:	289
Number:	16
Page Range:	S. 11262 - 11272
Date:	2014
Publisher:	AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
Place of Publication:	ROCKVILLE
ISSN:	1083-351X
Language:	English
Faculty:	Unspecified
Divisions:	Unspecified
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language RESISTANT NEPHROTIC SYNDROME; MULTIPLE SEQUENCE ALIGNMENT; GLOMERULAR SLIT DIAPHRAGM; TURKISH CHILDREN; N-GLYCOSYLATION; NPHS2 MUTATIONS; NEPHRIN; MEC-2; TRPC6; GENE Multiple languages Biochemistry & Molecular Biology Multiple languages
URI:	http://kups.ub.uni-koeln.de/id/eprint/44060