In the neuroepithelial lineage, reduced megalin protein levels were observed in DBS patient cells by day 9 of differentiation, compared to the control

In the neuroepithelial lineage, reduced megalin protein levels were observed in DBS patient cells by day 9 of differentiation, compared to the control. Rabbit Polyclonal to Cytochrome P450 2D6 tubule dysfunctions are typically encountered in congenital disorders due to defective endocytic receptors. DonnaiCBarrow or facioCoculo-acousticoCrenal syndrome (MIM no. 222448; herein referred as to DBS) is usually a rare inherited MLT-747 disease characterized by severe and multisystemic manifestations that include common craniofacial anomalies, highCgrade myopia, deafness and lowCmolecularCweight proteinuria. DBS is usually caused by lossCofCfunction mutations in the gene that encodes a protein called megalin C the most ancient member of the lowCdensity lipoprotein (LDL) receptor family (3). Being structurally related to a group of integral cell surface proteins, megalin is composed of a large extracellular domain name with several ligandCbinding regions, a transmembrane domain name and a short cytoplasmic tail with potential signalCactivation sequences. Compelling evidence suggests that megalin coordinates cellCfate trajectories, and hence cells behaviour and identity during brain development. This occurs through endocytosisCmediated uptake and control of the signalling of key differentiation factors, such as sonic hedgehog (SHH; ref. 3). In the proximal tubule of the kidney, megalin, in tandem with its coCreceptor cubilin and other accessory proteins, sustains the reabsorption and sorting of many essential nutrients within endolysosomal organelles for their subsequent degradation and recycling (3). Loss or mutation of megalin hinders crucial developmental processes and disrupts the physiology of kidney tubule, ultimately causing devastating malformations and severe life threateningCcomplications in humans (2,3). Mechanistically, how diseaseCcausing mutations trigger megalin dysfunction remains poorly comprehended. Assessments of the pathogenicity of putative diseaseCcausing variants require studies in physiologically relevant cell culture basedCsystems. MLT-747 For megalin, main epithelial cells are the most relevant system; however, these cells are shortClived and can only be procured from internal organs such as the kidneys, substantially limiting acquisition from individuals with rare disease causingCmutations. Now a study (4) by Flemming and colleagues in this issue of demonstrates a way of overcoming this hurdle. The experts took advantage of the recent improvements in induced pluripotent stem cell (iPSC) technology (5) to produce human disease in a dish models. Based on the elegant use of patientCderived iPSC cell lines, the authors deciphered the cascade of molecular events driving the dysfunction of the receptor megalin pathway in a family affected with DBS. First, the authors performed wholeCexome sequencing studies to identify putative genetic variants that are causal for the disease in the two affected family members. The sequencing analyses flagged a previously unknown homozygosity for any novel missense mutation that results in amino acid alteration in an epidermal growth factor (EGF) Ctype repeat in the extracellular domain name of megalin (LRP2R3192Q). This prospects in turn to deficiency of megalin and hence to kidney tubule dysfunction, as documented by expression studies showing absence of megalin patterning from your brush border of proximal tubular cells in renal biopsies and by the metabolite signature of proximal tubule dysfunction and renal Fanconi syndrome (i.e. hypercalciuria and low molecular excess weight proteinuria) encountered in the urine of both patients. Having recognized the diseaseCcausing variant, the authors proceeded to derive human iPSCs by reprogramming blood cells from these two affected siblings. Because of their intrinsic properties of indefinite selfCrenewal and potential to adopt virtually any cellular fate through differentiation, patientCspecific iPSCs represent a valuable tool for the study of disease mechanisms, as they can recapitulate complex features of MLT-747 the disease phenotype while retaining the convenience of systems. The authors applied established directed differentiation protocols to convert patientCspecific iPSCs derivates into physiologically relevant cell types that are affected in the disease and were previously inaccessible. To generate neurons, a well-established differentiation protocol was applied, in which iPS cells were cultured in neuronal growth.