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Congenital anomalies of the kidney and urinary tract (CAKUT) are the most common cause of chronic kidney disease (~ 45%) that manifests before 30 years of age. The genetic locus containing COL4A1 (13q33–34) has been implicated in vesicoureteral reflux (VUR), but mutations in COL4A1 have not been reported in CAKUT. We hypothesized that COL4A1 mutations cause CAKUT in humans. We performed whole exome sequencing (WES) in 550 families with CAKUT. As negative control cohorts we used WES sequencing data from patients with nephronophthisis (NPHP) with no genetic cause identified (n = 257) and with nephrotic syndrome (NS) due to monogenic causes (n = 100). We identified a not previously reported heterozygous missense variant in COL4A1 in three siblings with isolated VUR. When examining 549 families with CAKUT, we identified nine additional different heterozygous missense mutations in COL4A1 in 11 individuals from 11 unrelated families with CAKUT, while no COL4A1 mutations were identified in a control cohort with NPHP and only one in the cohort with NS. Most individuals (12/14) had isolated CAKUT with no extrarenal features. The predominant phenotype was VUR (9/14). There were no clinical features of the COL4A1-related disorders (e.g., HANAC syndrome, porencephaly, tortuosity of retinal arteries). Whereas COL4A1-related disorders are typically caused by glycine substitutions in the collagenous domain (84.4% of variants), only one variant in our cohort is a glycine substitution within the collagenous domain (1/10). We identified heterozygous COL4A1 mutations as a potential novel autosomal dominant cause of CAKUT that is allelic to the established COL4A1-related disorders and predominantly caused by non-glycine substitutions. [ABSTRACT FROM AUTHOR]

Mosaicism due to somatic mutations can cause multiple diseases including cancer, developmental and overgrowth syndromes, neurodevelopmental disorders, autoinflammatory diseases, and atrial fibrillation. With the increased use of next generation sequencing technology, multiple tools have been developed to identify low-frequency variants, specifically from matched tumor-normal tissues in cancer studies. To investigate whether mosaic variants are implicated in congenital heart disease (CHD), we developed a pipeline using the cancer somatic variant caller MuTect to identify mosaic variants in whole-exome sequencing (WES) data from a cohort of parent/affected child trios (n = 715) and a cohort of healthy individuals (n = 416). This is a novel application of the somatic variant caller designed for cancer to WES trio data. We identified two cases with mosaic KMT2D mutations that are likely pathogenic for CHD, but conclude that, overall, mosaicism detectable in peripheral blood or saliva does not account for a significant portion of CHD etiology. [ABSTRACT FROM AUTHOR]

We report on whole-exome sequencing (WES) of 213 melanomas. Our analysis established NF1, encoding a negative regulator of RAS, as the third most frequently mutated gene in melanoma, after BRAF and NRAS. Inactivating NF1 mutations were present in 46% of melanomas expressing wild-type BRAF and RAS, occurred in older patients and showed a distinct pattern of co-mutation with other RASopathy genes, particularly RASA2. Functional studies showed that NF1 suppression led to increased RAS activation in most, but not all, melanoma cases. In addition, loss of NF1 did not predict sensitivity to MEK or ERK inhibitors. The rebound pathway, as seen by the induction of phosphorylated MEK, occurred in cells both sensitive and resistant to the studied drugs. We conclude that NF1 is a key tumor suppressor lost in melanomas, and that concurrent RASopathy gene mutations may enhance its role in melanomagenesis. [ABSTRACT FROM AUTHOR]

Cutaneous T cell lymphoma (CTCL) is a non-Hodgkin lymphoma of skin-homing T lymphocytes. We performed exome and whole-genome DNA sequencing and RNA sequencing on purified CTCL and matched normal cells. The results implicate mutations in 17 genes in CTCL pathogenesis, including genes involved in T cell activation and apoptosis, NF-κB signaling, chromatin remodeling and DNA damage response. CTCL is distinctive in that somatic copy number variants (SCNVs) comprise 92% of all driver mutations (mean of 11.8 pathogenic SCNVs versus 1.0 somatic single-nucleotide variant per CTCL). These findings have implications for new therapeutics. [ABSTRACT FROM AUTHOR]

Upon detection of pathogen-associated molecular patterns, innate immune receptors initiate inflammatory responses. These receptors include cytoplasmic NOD-like receptors (NLRs) whose stimulation recruits and proteolytically activates caspase-1 within the inflammasome, a multiprotein complex. Caspase-1 mediates the production of interleukin-1 family cytokines (IL1FCs), leading to fever and inflammatory cell death (pyroptosis). Mutations that constitutively activate these pathways underlie several autoinflammatory diseases with diverse clinical features. We describe a family with a previously unreported syndrome featuring neonatal-onset enterocolitis, periodic fever, and fatal or near-fatal episodes of autoinflammation. We show that the disease is caused by a de novo gain-of-function mutation in NLRC4 encoding a p.Val341Ala substitution in the HD1 domain of the protein that cosegregates with disease. Mutant NLRC4 causes constitutive IL1FC production and macrophage cell death. Infected macrophages from affected individuals are polarized toward pyroptosis and exhibit abnormal staining for inflammasome components. These findings identify and describe the cause of a life-threatening but treatable autoinflammatory disease that underscores the divergent roles of the NLRC4 inflammasome. [ABSTRACT FROM AUTHOR]

Adrenal tumors autonomously producing cortisol cause Cushing's syndrome. We performed exome sequencing of 25 tumor-normal pairs and identified 2 subgroups. Eight tumors (including three carcinomas) had many somatic copy number variants (CNVs) with frequent deletion of CDC42 and CDKN2A, amplification of 5q31.2 and protein-altering mutations in TP53 and RB1. Seventeen tumors (all adenomas) had no somatic CNVs or TP53 or RB1 mutations. Six of these had known gain-of-function mutations in CTNNB1 (β-catenin) or GNAS (Gαs). Six others had somatic mutations in PRKACA (protein kinase A (PKA) catalytic subunit) resulting in a p.Leu206Arg substitution. Further sequencing identified this mutation in 13 of 63 tumors (35% of adenomas with overt Cushing's syndrome). PRKACA, GNAS and CTNNB1 mutations were mutually exclusive. Leu206 directly interacts with the regulatory subunit of PKA, PRKAR1A. Leu206Arg PRKACA loses PRKAR1A binding, increasing the phosphorylation of downstream targets. PKA activity induces cortisol production and cell proliferation, providing a mechanism for tumor development. These findings define distinct mechanisms underlying adrenal cortisol-producing tumors. [ABSTRACT FROM AUTHOR]

A cost-efficient way to increase power in a genetic association study is to pool controls from different sources. The genotyping effort can then be directed to large case series. The Nordic Control database, NordicDB, has been set up as a unique resource in the Nordic area and the data are available for authorized users through the web portal (http://www.nordicdb.org). The current version of NordicDB pools together high-density genome-wide SNP information from ∼5000 controls originating from Finnish, Swedish and Danish studies and shows country-specific allele frequencies for SNP markers. The genetic homogeneity of the samples was investigated using multidimensional scaling (MDS) analysis and pairwise allele frequency differences between the studies. The plot of the first two MDS components showed excellent resemblance to the geographical placement of the samples, with a clear NW-SE gradient. We advise researchers to assess the impact of population structure when incorporating NordicDB controls in association studies. This harmonized Nordic database presents a unique genome-wide resource for future genetic association studies in the Nordic countries. [ABSTRACT FROM AUTHOR]

B cells, αβ T cells and γδ T cells are conserved lymphocyte subtypes encoding their antigen receptors from somatically rearranged genes. αβ T cells undergo positive selection in the thymus by engagement of their T cell receptors (TCRs) with self-peptides presented by major histocompatibility complex molecules. The molecules that select γδ T cells are unknown. Vγ5+Vδ1+ cells comprise 90% of mouse epidermal γδ T cells. By mapping and genetic complementation using a strain showing loss of Vγ5+Vδ1+ cells due to a failure of thymic selection, we show that this defect is caused by mutation in Skint1, a newly identified gene expressed in thymus and skin that encodes a protein with immunoglobulin-like and transmembrane domains. Skint1 is the prototypic member of a rapidly evolving family of at least 11 genes in mouse, with greatest similarity to the butyrophilin genes. These findings define a new family of proteins mediating key epithelial-immune interactions. [ABSTRACT FROM AUTHOR]

The effects of alleles in many genes are believed to contribute to common complex diseases such as hypertension. Whether risk alleles comprise a small number of common variants or many rare independent mutations at trait loci is largely unknown. We screened members of the Framingham Heart Study (FHS) for variation in three genes—SLC12A3 (NCCT), SLC12A1 (NKCC2) and KCNJ1 (ROMK)—causing rare recessive diseases featuring large reductions in blood pressure. Using comparative genomics, genetics and biochemistry, we identified subjects with mutations proven or inferred to be functional. These mutations, all heterozygous and rare, produce clinically significant blood pressure reduction and protect from development of hypertension. Our findings implicate many rare alleles that alter renal salt handling in blood pressure variation in the general population, and identify alleles with health benefit that are nonetheless under purifying selection. These findings have implications for the genetic architecture of hypertension and other common complex traits. [ABSTRACT FROM AUTHOR]

Familial hypomagnesemic hypercalciuria and nephrocalcinosis (FHHNC [MIM 248250]) is a rare renal tubular disorder characterized by impaired reabsorption of magnesium and calcium in the thick ascending limb of Henle’s loop (tALH), causing renal magnesium wasting and hypercalciuria. Patients with FHHNC usually present with recurrent urinary tract infections, polyuria, nephrolithiasis (NL) and nephrocalcinosis (NC) with many progressing to chronic renal failure (CRF). We have shown recently that loss of function mutations in paracellin-1 PCLN-1/claudin-16, a renal tight junction protein located in the TAL, are causative of FHHNC. We present clinical and molecular studies on a highly inbred family with FHHNC in association with an unusual phenotype in that all affected members were extremely short. Affected individuals were found to be homozygous for marker D3S1314 on chromosome 3q. Sequencing of the PCLN-1/claudin-16 gene revealed a previously unknown point mutation at S235Y on exon 4 on the 4th transmembrane domain, providing additional evidence that inactivating mutations in the PCLN-1/claudin-16 gene result in FHHNC. [ABSTRACT FROM AUTHOR]

The mechanisms that govern homeostasis of complex systems have been elusive but can be illuminated by mutations that disrupt system behavior. Mutations in the gene encoding the kinase WNK4 cause pseudohypoaldosteronism type II (PHAII), a syndrome featuring hypertension and hyperkalemia. We show that physiology in mice transgenic for genomic segments harboring wild-type (TgWnk4WT) or PHAII mutant (TgWnk4PHAII) Wnk4 is changed in opposite directions: TgWnk4PHAII mice have higher blood pressure, hyperkalemia, hypercalciuria and marked hyperplasia of the distal convoluted tubule (DCT), whereas the opposite is true in TgWnk4WT mice. Genetic deficiency for the Na-Cl cotransporter of the DCT (NCC) reverses phenotypes seen in TgWnk4PHAII mice, demonstrating that the effects of the PHAII mutation are due to altered NCC activity. These findings establish that Wnk4 is a molecular switch that regulates the balance between NaCl reabsorption and K+ secretion by altering the mass and function of the DCT through its effect on NCC. [ABSTRACT FROM AUTHOR]

Whole-genome association studies are predicted to be especially powerful in isolated populations owing to increased linkage disequilibrium (LD) and decreased allelic diversity, but this possibility has not been empirically tested. We compared genome-wide data on 113,240 SNPs typed on 30 trios from the Pacific island of Kosrae to the same markers typed in the 270 samples from the International HapMap Project. The extent of LD is longer and haplotype diversity is lower in Kosrae than in the HapMap populations. More than 98% of Kosraen haplotypes are present in HapMap populations, indicating that HapMap will be useful for genetic studies on Kosrae. The long-range LD around common alleles and limited diversity result in improved efficiency in genetic studies in this population and augments the power to detect association of 'hidden SNPs'. [ABSTRACT FROM AUTHOR]

A key question in systems biology is how diverse physiologic processes are integrated to produce global homeostasis. Genetic analysis can contribute by identifying genes that perturb this integration. One system orchestrates renal NaCl and K+ flux to achieve homeostasis of blood pressure and serum K+ concentration (refs. 2,3). Positional cloning implicated the serine-threonine kinase WNK4 in this process; clustered mutations in PRKWNK4, encoding WNK4, cause hypertension and hyperkalemia (pseudohypoaldosteronism type II, PHAII) by altering renal NaCl and K+ handling. Wild-type WNK4 inhibits the renal Na-Cl cotransporter (NCCT); mutations that cause PHAII relieve this inhibition. This explains the hypertension of PHAII but does not account for the hyperkalemia. By expression in Xenopus laevis oocytes, we show that WNK4 also inhibits the renal K+ channel ROMK. This inhibition is independent of WNK4 kinase activity and is mediated by clathrin-dependent endocytosis of ROMK, mechanisms distinct from those that characterize WNK4 inhibition of NCCT. Most notably, the same mutations in PRKWNK4 that relieve NCCT inhibition markedly increase inhibition of ROMK. These findings establish WNK4 as a multifunctional regulator of diverse ion transporters; moreover, they explain the pathophysiology of PHAII. They also identify WNK4 as a molecular switch that can vary the balance between NaCl reabsorption and K+ secretion to maintain integrated homeostasis. [ABSTRACT FROM AUTHOR]

End-stage renal disease (ESRD) is a major public health problem, affecting 1 in 1,000 individuals and with an annual death rate of 20% despite dialysis treatment. IgA nephropathy (IgAN) is the most common form of glomerulonephritis, a principal cause of ESRD worldwide; it affects up to 1.3% of the population and its pathogenesis is unknown. Kidneys of people with IgAN show deposits of IgA-containing immune complexes with proliferation of the glomerular mesangium (Fig. 1). Typical clinical features include onset before age 40 with haematuria and proteinuria (blood and protein in the urine), and episodes of gross haematuria following mucosal infections are common; 30% of patients develop progressive renal failure. Although not generally considered a hereditary disease, striking ethnic variation in prevalence and familial clustering, along with subclinical renal abnormalities among relatives of IgAN cases, have suggested a heretofore undefined genetic component. By genome-wide analysis of linkage in 30 multiplex IgAN kindreds, we demonstrate linkage of IgAN to 6q22?23 under a dominant model of transmission with incomplete penetrance, with a lod score of 5.6 and 60% of kindreds linked. These findings for the first time indicate the existence of a locus with large effect on development of IgAN and identify the chromosomal location of this disease gene. [ABSTRACT FROM AUTHOR]

The multi-subunit H+-ATPase pump is present at particularly high density on the apical (luminal) surface of α-intercalated cells of the cortical collecting duct of the distal nephron, where vectorial proton transport is required for urinary acidification. The complete subunit composition of the apical ATPase, however, has not been fully agreed upon. Functional failure of α-intercalated cells results in a group of disorders, the distal renal tubular acidoses (dRTA), whose features include metabolic acidosis accompanied by disturbances of potassium balance, urinary calcium solubility, bone physiology and growth. Mutations in the gene encoding the B-subunit of the apical pump (ATP6B1) cause dRTA accompanied by deafness. We previously localized a gene for dRTA with preserved hearing to 7q33?34 (ref. 4). We report here the identification of this gene, ATP6N1B, which encodes an 840 amino acid novel kidney-specific isoform of ATP6N1A, the 116-kD non-catalytic accessory subunit of the proton pump. Northern-blot analysis demonstrated ATP6N1B expression in kidney but not other main organs. Immunofluorescence studies in human kidney cortex revealed that ATP6N1B localizes almost exclusively to the apical surface of α-intercalated cells. We screened nine dRTA kindreds with normal audiometry that linked to the ATP6N1B locus, and identified different homozygous mutations in ATP6N1B in eight. These include nonsense, deletion and splice-site changes, all of which will truncate the protein. Our findings identify a new kidney-specific proton pump 116-kD accessory subunit that is highly expressed in proton-secreting cells in the distal nephron, and illustrate its essential role in normal vectorial acid transport into the urine by the kidney. [ABSTRACT FROM AUTHOR]

In evaluating hypertensive children and adolescents, the etiological considerations should include a set of inherited disorders that share very low plasma renin activity (PRA) as a common feature. In particular among these disorders, glucocorticoid remediable aldosteronism (GRA) appears to be emerging as an important etiology of hypertension in the pediatric population. We report the evaluation of a 9-year-old Caucasian girl who presented with severe hypertension and a strong family history of early-onset hypertension. Her suppressed PRA, her family history, and her failure to respond to conventional antihypertensive therapy raised GRA as a potential etiology. The diagnosis was confirmed by an elevated ratio of urinary 18-oxotetrahydrocortisol to urinary tetrahydroaldosterone and genetic testing, which demonstrated the chimeric gene duplication. The molecular pathogenesis of GRA and the clinical implications are reviewed. [ABSTRACT FROM AUTHOR]

We describe a patient with signs and symptoms of classic Bartter syndrome. The patient tested negative for all known genetic abnormalities associated with this tubular disorder. Proteinuria was found within 1 year after the diagnosis of Bartter syndrome. A renal biopsy performed 6 months later, when her kidney function was normal, revealed focal segmental glomerulosclerosis (FSGS). We propose a link between stimulation of the renin-angiotensin system and sclerotic changes in the glomerulus. This lesion may explain previous reports of kidney failure in patients with Bartter syndrome. [ABSTRACT FROM AUTHOR]

Pseudohypoaldosteronism type I (PHA1) is characterized by neonatal renal salt wasting with dehydration, hypotension, hyperkalaemia and metabolic acidosis, despite elevated aldosterone levels. Two forms of PHA1 exist. An autosomal recessive form features severe disease with manifestations persisting into adulthood. This form is caused by loss-of-function mutations in genes encoding subunits of the amiloride-sensitive epithelial sodium channel (ENaC; refs 2,3). Autosomal dominant or sporadic PHA1 is a milder disease that remits with age. Among six dominant and seven sporadic PHA1 kindreds, we have found no ENaC gene mutations, implicating mutations in other genes. As ENaC activity in the kidney is regulated by the steroid hormone aldosterone acting through the mineralocorticoid receptor, we have screened the mineralocorticoid receptor gene (MLR) for variants and have identified heterozygous mutations in one sporadic and four dominant kindreds. These include two frameshift mutations (one a de novo mutation), two premature termination codons and one splice donor mutation. These mutations segregate with PHA1 and are not found in unaffected subjects. These findings demonstrate that heterozygous MLR mutations cause PHA1, underscore the important role of mineralocorticoid receptor function in regulation of salt and blood pressure homeostasis in humans and motivate further study of this gene for a potential role in blood pressure variation. [ABSTRACT FROM AUTHOR]

A correction to the article "Recurrent activating mutation in PRKACA in cortisol-producing adrenal tumors" that was published online in the April 20, 2014 issue is presented.

In the nervous system, the intracellular chloride concentration ([Cl[sup-]][subi]) determines the strength and polarity of γ-aminobutyric acid (GABA)-mediated neurotransmission. [CI[sup-]][subi] is determined, in part, by the activities of the SLC12 cation-chloride cotransporters (CCCs). These transporters include the Na-K-2C1 cotransporter NKCC1, which mediates chloride influx, and various K-Cl cotransporters — such as KCC2 and KCC3 — that extrude chloride. A precise balance between NKCC1 and KCC2 activity is necessary for inhibitory GABAergic signaling in the adult CNS, and for excitatory GABAergic signaling in the developing CNS and the adult PNS. Altered chloride homeostasis, resulting from mutation or dysfunction of NKCC1 and/or KCC2, causes neuronal hypoexcitability or hyperexcitabffity; such derangements have been implicated in the pathogenesis of seizures and neuropathic pain. [Cl[sup-]][subi] is also regulated to maintain normal cell volume. Dysfunction of NKCC1 or of swelling-activated K-Cl cotransporters has been implicated in the damaging secondary effects of cerebral edema after ischemic and traumatic brain injury, as well as in swelling-related neurodegeneration. CCCs represent attractive therapeutic targets in neurological disorders the pathogenesis of which involves deranged cellular chloride homoestasis. [ABSTRACT FROM AUTHOR]