Eric Olinger

The serine protease hepsin mediates urinary secretion and polymerisation of Zona Pellucida domain protein uromodulin

Uromodulin is the most abundant protein in the urine. It is exclusively produced by renal epithelial cells and it plays key roles in kidney function and disease. Uromodulin mainly exerts its function as an extracellular matrix whose assembly depends on a conserved, specific proteolytic cleavage leading to conformational activation of a Zona Pellucida (ZP) polymerisation domain. Through a comprehensive approach, including extensive characterisation of uromodulin processing in cellular models and in specific knock-out mice, we demonstrate that the membrane-bound serine protease hepsin is the enzyme responsible for the physiological cleavage of uromodulin. Our findings define a key aspect of uromodulin biology and identify the first in vivo substrate of hepsin. The identification of hepsin as the first protease involved in the release of a ZP domain protein is likely relevant for other members of this protein family, including several extracellular proteins, as egg coat proteins and inner ear tectorins. DOI: http://dx.doi.org/10.7554/eLife.08887.001

Parvalbumin: calcium and magnesium buffering in the distal nephron

Parvalbumin (PV) is a classical member of the EF-hand protein superfamily that has been described as a Ca(2+) buffer and Ca(2+) transporter/shuttle protein and may also play an additional role in Mg(2+) handling. PV is exclusively expressed in the early part of the distal convoluted tubule in the human and mouse kidneys. Recent studies in Pvalb knockout mice revealed a role of PV in the distal handling of electrolytes: the lack of PV was associated with a mild salt-losing phenotype with secondary aldosteronism, salt craving and stronger bones compared with controls. A link between the Ca(2+)-buffering capacity of PV and the expression of the thiazide-sensitive Na(+)-Cl(-) cotransporter was established, which could be relevant to the regulation of sodium transport in the distal nephron. Variants in the PVALB gene that encodes PV have been described, but their relevance to kidney function has not been established. PV is also considered a reliable marker of chromophobe carcinoma and oncocytoma, two neoplasms deriving from the distal nephron. The putative role of PV in tumour genesis remains to be investigated.

Uromodulin: from physiology to rare and complex kidney disorders

Uromodulin (also known as Tamm-Horsfall protein) is exclusively produced in the kidney and is the most abundant protein in normal urine. The function of uromodulin remains elusive, but the available data suggest that this protein might regulate salt transport, protect against urinary tract infection and kidney stones, and have roles in kidney injury and innate immunity. Interest in uromodulin was boosted by genetic studies that reported involvement of the UMOD gene, which encodes uromodulin, in a spectrum of rare and common kidney diseases. Rare mutations in UMOD cause autosomal dominant tubulointerstitial kidney disease (ADTKD), which leads to chronic kidney disease (CKD). Moreover, genome-wide association studies have identified common variants in UMOD that are strongly associated with risk of CKD and also with hypertension and kidney stones in the general population. These findings have opened up a new field of kidney research. In this Review we summarize biochemical, physiological, genetic and pathological insights into the roles of uromodulin; the mechanisms by which UMOD mutations cause ADTKD, and the association of common UMOD variants with complex disorders.

Tubular proteinuria in patients with HNF1α mutations: HNF1α drives endocytosis in the proximal tubule

Hepatocyte nuclear factor 1α (HNF1α) is a transcription factor expressed in the liver, pancreas, and proximal tubule of the kidney. Mutations of HNF1α cause an autosomal dominant form of diabetes mellitus (MODY-HNF1A) and tubular dysfunction. To gain insights into the role of HNF1α in the proximal tubule, we analyzed Hnf1a-deficient mice. Compared with wild-type littermates, Hnf1a knockout mice showed low-molecular-weight proteinuria and a 70% decrease in the uptake of β2-microglobulin, indicating a major endocytic defect due to decreased expression of megalin/cubilin receptors. We identified several binding sites for HNF1α in promoters of Lrp2 and Cubn genes encoding megalin and cubilin, respectively. The functional interaction of HNF1α with these promoters was shown in C33 epithelial cells lacking endogenous HNF1α. Defective receptor-mediated endocytosis was confirmed in proximal tubule cells from these knockout mice and could be rescued by transfection of wild-type but not mutant HNF1α. Transfection of human proximal tubule HK2 cells with HNF1α was able to upregulate megalin and cubilin expression and to increase endocytosis of albumin. Low-molecular-weight proteinuria was consistently detected in individuals with HNF1A mutations compared with healthy controls and patients with non-MODY-HNF1A diabetes mellitus. Thus, HNF1α plays a key role in the constitutive expression of megalin and cubilin, hence regulating endocytosis in the proximal tubule of the kidney. These findings provide new insight into the renal phenotype of individuals with mutations of HNF1A.

The value of ECG parameters as markers of treatment response in Fabry cardiomyopathy

Objective Best treatment outcomes in Fabry disease (FD) associated cardiomyopathy can be obtained when treatment is started as early as possible. The rationale of this study was to assess the role of ECG changes for identification of cardiac involvement and patients at an earlier stage of the disease more likely deriving a benefit from enzyme replacement therapy (ERT). Methods A retrospective analysis of patient data was performed from an observational, longitudinal, prospective cohort. Treatment response was defined as absence or presence of disease progression, defined as new onset or increase in left ventricular (LV) mass >10%. Demographic, clinical, ECG and echocardiographic parameters at baseline were tested for their value in determining absence or presence of disease progression under ERT at 5-year follow-up. Results The study population consisted of a total of 38 patients (25 men, mean age 36±13 years, overall median follow-up duration 6.4±1.2 years). Patients in the progression group (14 men, 4 women) had a longer QRS duration (99±11 ms vs 84±13 ms, p<0.05 for men, 93±9 years vs 81±5 years, p<0.05 for women) and QTc interval (401±15 ms vs 372±10 ms, p<0.005 for men) and a higher amount of ECG abnormalities (86% vs 18%, p<0.005 for men and 100% vs 0%, p<0.005 for women) at the time of ERT initiation. An abnormal baseline ECG was significantly associated with disease progression (sensitivity 94.1%, specificity 88.9%, positive likelihood ratio of 8.47, p<0.005). Conclusions An abnormal ECG at the time of treatment initiation is significantly associated with cardiac disease progression in FD. This effect seems to be independent of age, gender or LV mass at baseline and suggests maximal treatment benefit when ERT is initiated before ECG abnormalities develop.

Paradoxical response to furosemide in uromodulin-associated kidney disease

Mutations in the UMOD gene coding for uromodulin cause autosomal dominant tubulointerstitial kidney disease. Uromodulin is known to regulate transport processes in the thick ascending limb, but it remains unknown whether UMOD mutations are associated with functional tubular alterations in the early phase of the disease. The responses to furosemide and to a water deprivation test were compared in a 32-year-old female patient carrying the pathogenic UMOD mutation p.C217G and her unaffected 31-year-old sister. A single dose of furosemide induced an intense headache with exaggerated decrease in blood pressure (Δsyst: 30 versus 20 mmHg; Δdiast: 18 versus 5 mmHg) and body weight (Δ2.6 kg versus Δ0.9 kg over 3 h) in the proband versus unaffected sib. The diuretic response and the fall in urine osmolality were also more important and detected earlier in the affected sib. Water deprivation led to increased plasma osmolality and urine concentration in both siblings; however, the response to desmopressin was attenuated in the affected sib. These data reveal that mutations of uromodulin cause specific transport alterations, including exaggerated response to furosemide and a failure to maximally concentrate urine, in the early phase of the disease.

A mouse model for inherited renal fibrosis associated with endoplasmic reticulum stress

ABSTRACT Renal fibrosis is a common feature of renal failure resulting from multiple etiologies, including diabetic nephropathy, hypertension and inherited renal disorders. However, the mechanisms of renal fibrosis are incompletely understood and we therefore explored these by establishing a mouse model for a renal tubular disorder, referred to as autosomal dominant tubulointerstitial kidney disease (ADTKD) due to missense uromodulin (UMOD) mutations (ADTKD-UMOD). ADTKD-UMOD, which is associated with retention of mutant uromodulin in the endoplasmic reticulum (ER) of renal thick ascending limb cells, is characterized by hyperuricemia, interstitial fibrosis, inflammation and renal failure, and we used targeted homologous recombination to generate a knock-in mouse model with an ADTKD-causing missense cysteine to arginine uromodulin mutation (C125R). Heterozygous and homozygous mutant mice developed reduced uric acid excretion, renal fibrosis, immune cell infiltration and progressive renal failure, with decreased maturation and excretion of uromodulin, due to its retention in the ER. The ER stress marker 78 kDa glucose-regulated protein (GRP78) was elevated in cells expressing mutant uromodulin in heterozygous and homozygous mutant mice, and this was accompanied, both in vivo and ex vivo, by upregulation of two unfolded protein response pathways in primary thick ascending limb cells from homozygous mutant mice. However, this did not lead to an increase in apoptosis in vivo. Thus, we have developed a novel mouse model for renal fibrosis, which will be a valuable resource to decipher the mechanisms linking uromodulin mutations with ER stress and renal fibrosis. Summary: A mouse model for renal fibrosis caused by uromodulin mutations reveals roles for ER stress and the unfolded protein response.

Multiplex epithelium dysfunction due to CLDN10 mutation: the HELIX syndrome

PurposeWe aimed to identify the genetic cause to a clinical syndrome encompassing hypohidrosis, electrolyte imbalance, lacrimal gland dysfunction, ichthyosis, and xerostomia (HELIX syndrome), and to comprehensively delineate the phenotype.MethodsWe performed homozygosity mapping, whole-genome sequencing, gene sequencing, expression studies, functional tests, protein bioinformatics, and histological characterization in two unrelated families with HELIX syndrome.ResultsWe identified biallelic missense mutations (c.386C>T, p.S131L and c.2T>C, p.M1T) in CLDN10B in six patients from two unrelated families. CLDN10B encodes Claudin-10b, an integral tight junction (TJ) membrane-spanning protein expressed in the kidney, skin, and salivary glands. All patients had hypohidrosis, renal loss of NaCl with secondary hyperaldosteronism and hypokalemia, as well as hypolacrymia, ichthyosis, xerostomia, and severe enamel wear. Functional testing revealed that patients had a decreased NaCl absorption in the thick ascending limb of the loop of Henle and a severely decreased secretion of saliva. Both mutations resulted in reduced or absent Claudin-10 at the plasma membrane of epithelial cells.ConclusionCLDN10 mutations cause a dysfunction in TJs in several tissues and, subsequently, abnormalities in renal ion transport, ectodermal gland homeostasis, and epidermal integrity.

Claudins: a tale of interactions in the thick ascending limb

The claudins are the main proteins composing the tight junctions. The differential expression of claudin isoforms contributes to the specificity of paracellular transport pathways along the kidney tubule. Recent studies focusing on claudin-10 and claudin-16 support the longitudinal specialization of paracellular transport of cations within the thick ascending limb and demonstrate that complex tubular adaptations operate to buffer the consequences of chronic segmental lesions in the kidney.

A novel homozygous UMOD mutation reveals gene dosage effects on uromodulin processing and urinary excretion

Abstract Heterozygous mutations in UMOD encoding the urinary protein uromodulin are the most common genetic cause of autosomal dominant tubulointerstitial kidney disease (ADTKD). We describe the exceptional case of a patient from a consanguineous family carrying a novel homozygous UMOD mutation (p.C120Y) affecting a conserved cysteine residue within the EGF-like domain III of uromodulin. Comparison of heterozygote and homozygote mutation carriers revealed a gene dosage effect with unprecedented low levels of uromodulin and aberrant uromodulin fragments in the urine of the homozygote proband. Despite an amplified biological effect of the homozygote mutation, the proband did not show a strikingly more severe clinical evolution nor was the near absence of urinary uromodulin associated with urinary tract infections or kidney stones.