Co-inheritance of a PKD1 mutation and homozygous PKD2 variant: A potential modifier in autosomal dominant polycystic kidney disease

Background: Autosomal dominant polycystic kidney disease (ADPKD), which is caused by mutations in polycystins 1 (PC1) and 2 (PC2), is one of the most commonly inherited renal diseases, affecting ∼1 : 1000 Caucasians. Materials and methods: We screened Greek ADPKD patients with the denaturing gradient gel electrophoresis (DGGE) assay and direct sequencing. Results: We identified a patient homozygous for a nucleotide change c.1445T > G, resulting in a novel homozygous substitution of the non-polar hydrophobic phenylalanine to the polar hydrophilic cysteine in exon 6 at codon 482 (p.F482C) of the PKD2 gene and a de-novo PKD1 splice-site variant IVS21-2delAG. We did not find this PKD2 variant in a screen of 280 chromosomes of healthy subjects, supporting its pathogenicity. The proband's parents did not have the PKD1 mutation. Real-time PCR of the PKD2 transcript from a skin biopsy revealed 20-fold higher expression in the patient than in a healthy subject and was higher in the patient's peripheral blood mononuclear cells (PBMCs) than in those of her heterozygote daughter and a healthy subject. The greater gene expression was also supported by Western blotting. Inner medullar collecting duct (IMCD) cells transfected with the mutant PKD2 mouse gene presented a perinuclear and diffuse cytoplasmic localization compared with the wild type ER localization. Patch-clamping of PBMCs from the p.F482C homozygous and heterozygous subjects revealed lower polycystin-2 channel function than in controls. Conclusions: We report for the first time a patient with ADPKD who is heterozygous for a de novo PKD1 variant and homozygous for a novel PKD2 mutation. © 2008 The Authors

Regulation of Sodium Channel Activity by Capping of Actin Filaments

Ion transport in various tissues can be regulated by the cortical actin cytoskeleton. Specifically, involvement of actin dynamics in the regulation of nonvoltage-gated sodium channels has been shown. Herein, inside-out patch clamp experiments were performed to study the effect of the heterodimeric actin capping protein CapZ on sodium channel regulation in leukemia K562 cells. The channels were activated by cytochalasin-induced disruption of actin filaments and inactivated by G-actin under ionic conditions promoting rapid actin polymerization. CapZ had no direct effect on channel activity. However, being added together with G-actin, CapZ prevented actin-induced channel inactivation, and this effect occurred at CapZ/actin molar ratios from 1:5 to 1:100. When actin was allowed to polymerize at the plasma membrane to induce partial channel inactivation, subsequent addition of CapZ restored the channel activity. These results can be explained by CapZ-induced inhibition of further assembly of actin filaments at the plasma membrane due to the modification of actin dynamics by CapZ. No effect on the channel activity was observed in response to F-actin, confirming that the mechanism of channel inactivation does not involve interaction of the channel with preformed filaments. Our data show that actin-capping protein can participate in the cytoskeleton-associated regulation of sodium transport in nonexcitable cells