G. M. Ghiggeri

Mutation of the Mg2+ Transporter SLC41A1 Results in a Nephronophthisis-Like Phenotype

Nephronophthisis (NPHP)-related ciliopathies are recessive, single-gene disorders that collectively make up the most common genetic cause of CKD in the first three decades of life. Mutations in 1 of the 15 known NPHP genes explain less than half of all cases with this phenotype, however, and the recently identified genetic causes are exceedingly rare. As a result, a strategy to identify single-gene causes of NPHP-related ciliopathies in single affected families is needed. Although whole-exome resequencing facilitates the identification of disease genes, the large number of detected genetic variants hampers its use. Here, we overcome this limitation by combining homozygosity mapping with whole-exome resequencing in a sibling pair with an NPHP-related ciliopathy. Whole-exome capture revealed a homozygous splice acceptor site mutation (c.698G>T) in the renal Mg(2+) transporter SLC41A1. This mutation resulted in skipping of exon 6 of SLC41A1, resulting in an in-frame deletion of a transmembrane helix. Transfection of cells with wild-type or mutant SLC41A1 revealed that deletion of exon 6 completely blocks the Mg(2+) transport function of SLC41A1. Furthermore, in normal human kidney tissue, endogenous SLC41A1 specifically localized to renal tubules situated at the corticomedullary boundary, consistent with the region of cystogenesis observed in NPHP and related ciliopathies. Last, morpholino-mediated knockdown of slc41a1 expression in zebrafish resulted in ventral body curvature, hydrocephalus, and cystic kidneys, similar to the effects of knocking down other NPHP genes. Taken together, these data suggest that defects in the maintenance of renal Mg(2+) homeostasis may lead to tubular defects that result in a phenotype similar to NPHP.

Mapping of the human COL5A1 gene to chromosome 9q34.3

SummaryA 353-bp region encoding for the NH2 terminus of the noncollagenic part of the αl(V) chain was amplified by the polymerase chain reaction (PCR), subcloned and sequenced. The subcloned PCR product (pGC1) presented the same nucleotide sequence as the original fragment from the published sequence of COL5A1. In situ hybridization, using pGC1 as a probe, mapped the COL5A1 gene to chromosome 9q34.3. This assignement shows that COL5A1 is not synthenic with COL5A2, which is localized together with other collagen genes on chromosome 2.

Are the nail-patella syndrome and the autosomal Goltz-like syndrome the phenotypic expressions of different alleles at the COL5A1 locus ?

The COL5A1 gene, which encodes the pro α1(V) chain, was recently mapped to 9q34.3 in the same region as the nail-patella locus. This was taken as an indication that the nail-patella syndrome may be an inherited connective tissue disorder. We demonstrate COL5A1 heterozygous deletion and fibroblast under-expression of α1(V) chains in a girl with an unbalanced translocation resulting in 9q32→qter monosomy. The patient presents dysplastic nails, a sign typical of nail-patella syndrome, but normal patella. Moreover, she has skin and bone disorders similar to those found in the Goltz syndrome. We suggest that monosomy for the COL5A1 gene is responsible for these connective tissue disorders. Accordingly, the nail-patella syndrome could be attributable to mutations inside the COL5A1 gene rather than to a deletion of it.

Molecular and Cellular Mechanisms for Proteinuria in Minimal Change Disease

Minimal Change Disease (MCD) is a clinical condition characterized by acute nephrotic syndrome, no evident renal lesions at histology and good response to steroids. However, frequent recurrence of the disease requires additional therapies associated with steroids. Such multi-drug dependence and frequent relapses may cause disease evolution to focal and segmental glomerulosclerosis (FSGS) over time. The differences between the two conditions are not well defined, since molecular mechanisms may be shared by the two diseases. In some cases, genetic analysis can make it possible to distinguish MCD from FSGS; however, there are cases of overlap. Several hypotheses on mechanisms underlying MCD and potential molecular triggers have been proposed. Most studies were conducted on animal models of proteinuria that partially mimic MCD and may be useful to study glomerulosclerosis evolution; however, they do not demonstrate a clear-cut separation between MCD and FSGS. Puromycin Aminonucleoside and Adriamycin nephrosis are models of glomerular oxidative damage, characterized by loss of glomerular basement membrane polyanions resembling MCD at the onset and, at more advanced stages, by glomerulosclerosis resembling FSGS. Also Buffalo/Mna rats present initial lesions of MCD, subsequently evolving to FSGS; this mechanism of renal damage is clearer since this rat strain inherits the unique characteristic of overexpressing Th2 cytokines. In Lipopolysaccharide nephropathy, an immunological condition of renal toxicity linked to B7-1(CD80), mice develop transient proteinuria that lasts a few days. Overall, animal models are useful and necessary considering that they reproduce the evolution from MCD to FSGS that is, in part, due to persistence of proteinuria. The role of T/Treg/Bcells on human MCD has been discussed. Many cytokines, immunomodulatory mechanisms, and several molecules have been defined as a specific cause of proteinuria. However, the hypothesis of a single cell subset or molecule as cause of MCD is not supported by research and an interactive process seems more logical. The implication or interactive role of oxidants, Th2 cytokines, Th17, Tregs, B7-1(CD80), CD40/CD40L, c-Mip, TNF, uPA/suPAR, Angiopoietin-like 4 still awaits a definitive confirmation. Whole genome sequencing studies could help to define specific genetic features that justify a definition of MCD as a “clinical-pathology-genetic entity.”