title: Uromodulin mutations causing familial juvenile hyperuricaemic nephropathy lead to protein maturation defects and retention in the endoplasmic reticulum.
abstract: Familial juvenile hyperuricaemic nephropathy (FJHN), an autosomal dominant disorder, is caused by mutations in the UMOD gene, which encodes Uromodulin, a glycosylphosphatidylinositol-anchored protein that is expressed in the thick ascending limb of the loop of Henle and excreted in the urine. Uromodulin contains three epidermal growth factor (EGF)-like domains, a cysteine-rich region which includes a domain of eight cysteines and a zona pellucida (ZP) domain. Over 90% of UMOD mutations are missense, and 62% alter a cysteine residue, implicating a role for protein misfolding in the disease. We investigated 20 northern European FJHN probands for UMOD mutations. Wild-type and mutant Uromodulins were functionally studied by expression in HeLa cells and by the use of western blot analysis and confocal microscopy. Six different UMOD missense mutations (Cys32Trp, Arg185Gly, Asp196Asn, Cys217Trp, Cys223Arg and Gly488Arg) were identified. Patients with UMOD mutations were phenotypically similar to those without UMOD mutations. The mutant Uromodulins had significantly delayed maturation, retention in the endoplasmic reticulum (ER) and reduced expression at the plasma membrane. However, Gly488Arg, which is the only mutation we identified in the ZP domain, was found to be associated with milder in vitro abnormalities and to be the only mutant Uromodulin detected in conditioned medium from transfected cells, indicating that the severity of the mutant phenotypes may depend on their location within the protein. Thus, FJHN-causing Uromodulin mutants are retained in the ER, with impaired intracellular maturation and trafficking, thereby indicating mechanisms whereby Uromodulin mutants may cause the phenotype of FJHN.
authors.affiliation: Academic Endocrine Unit, Churchill Hospital, Oxford Centre for Diabetes, Endocrinology and Metabolism, University of Oxford, Headington, Oxford OX3 7LJ, UK.
authors.forename: Siân E
authors.initials: SE
authors.lastname: Williams
authors.forename: Anita A C
authors.initials: AA
authors.lastname: Reed
authors.forename: Juris
authors.initials: J
authors.lastname: Galvanovskis
authors.forename: Corinne
authors.initials: C
authors.lastname: Antignac
authors.forename: Tim
authors.initials: T
authors.lastname: Goodship
authors.forename: Fiona E
authors.initials: FE
authors.lastname: Karet
authors.forename: Peter
authors.initials: P
authors.lastname: Kotanko
authors.forename: Karl
authors.initials: K
authors.lastname: Lhotta
authors.forename: Vincent
authors.initials: V
authors.lastname: Morinière
authors.forename: Paul
authors.initials: P
authors.lastname: Williams
authors.forename: William
authors.initials: W
authors.lastname: Wong
authors.forename: Patrik
authors.initials: P
authors.lastname: Rorsman
authors.forename: Rajesh V
authors.initials: RV
authors.lastname: Thakker
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D004721:Endoplasmic Reticulum / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005260:Female; D006367:HeLa Cells; D006801:Humans; D033461:Hyperuricemia / Q000235:genetics* / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D009088:Mucoproteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D020125:Mutation, Missense; D010375:Pedigree; D017510:Protein Folding; D021381:Protein Transport; D058949:Uromodulin; D044465:White People / Q000235:genetics; D055815:Young Adult
chemical_list: D009088:Mucoproteins; C548612:UMOD protein, human; D058949:Uromodulin