The chloride intracellular channel 5A (CLIC5A) protein, one of two isoforms produced by the CLIC5 gene, was isolated originally as part of a cytoskeletal protein complex containing ezrin from placental microvilli. Whether CLIC5A functions as a bona fide ion channel is controversial. We reported previously that a CLIC5 transcript is enriched approximately 800-fold in human renal glomeruli relative to most other tissues. Therefore, this study sought to explore CLIC5 expression and function in glomeruli. RT-PCR and Western blots show that CLIC5A is the predominant CLIC5 isoform expressed in glomeruli. Confocal immunofluorescence and immunogold electron microscopy reveal high levels of CLIC5A protein in glomerular endothelial cells and podocytes. In podocytes, CLIC5A localizes to the apical plasma membrane of foot processes, similar to the known distribution of podocalyxin and ezrin. Ezrin and podocalyxin colocalize with CLIC5A in glomeruli, and podocalyxin coimmunoprecipitates with CLIC5A from glomerular lysates. In glomeruli of jitterbug (jbg/jbg) mice, which lack the CLIC5A protein, ezrin and phospho-ERM levels in podocytes are markedly lower than in wild-type mice. Transmission electron microscopy reveals patchy broadening and effacement of podocyte foot processes as well as vacuolization of glomerular endothelial cells. These ultrastructural changes are associated with microalbuminuria at baseline and increased susceptibility to adriamycin-induced glomerular injury compared with wild-type mice. Together, the data suggest that CLIC5A is required for the development and/or maintenance of the proper glomerular endothelial cell and podocyte architecture. We postulate that the interaction between podocalyxin and subjacent filamentous actin, which requires ezrin, is compromised in podocytes of CLIC5A-deficient mice, leading to dysfunction under unfavorable genetic or environmental conditions.

Axenfeld-Rieger syndrome (ARS) and iridogoniodysgenesis syndrome (IGDS) are clinically related autosomal dominant disorders which affect the anterior segment of the eye as well as non-ocular structures. ARS patients present with iris hypoplasia, a prominent Schwalbe line, adhesions between the iris stroma and the iridocorneal angle and increased intraocular pressure. IGDS is characterized by iris hypoplasia, goniodysgenesis and increased intraocular pressure. Each syndrome also presents with non-ocular features including maxillary hypoplasia, micro and anodontia, redundant periumbilical skin, hypospadius (in males), and each has been genetically linked to chromosome 4q25. RIEG1 , the gene responsible for the 4q25 ARS phenotype, recently has been cloned. RIEG1 encodes a novel member of the bicoid class of homeobox proteins known to be active as transcription factors. Mutational analysis has previously detected several mutations in this gene in ARS individuals. We have now detected a mutation in RIEG1 which segregates with the disease phenotype in a family with IGDS. This mutation is a G-->A transition altering an arginine residue to a histidine in a highly conserved location in the second helix of the homeobox of RIEG1. This mutation indicates that IGDS and ARS are allelic variants of the same disorder. This wide variability in clinical consequences of mutations at the RIEG1 4q25 locus implicates the RIEG gene broadly in ocular and craniofacial disorders.

SummaryPlakoglobin (c-catenin), a constituent of the adherens junction and desmosomes, has signaling capabilities typically associated with tumor/metastasis suppression through mechanisms that remain undefined. To determine the role of plakoglobin during tumorigenesis and metastasis, we expressed plakoglobin in human tongue squamous cell carcinoma (SCC9) cells and compared the mRNA profiles of parental SCC9 cells and their plakoglobin-expressing transfectants (SCC9-PG). We detected several p53-target genes whose levels were altered upon plakoglobin expression. In this study, we identified the p53 regulated tumor suppressor 14-3-3s as a direct plakoglobin-p53 target gene. Coimmunoprecipitation experiments revealed that plakoglobin and p53 interact, and chromatin immunoprecipitation and electrophoretic mobility shift assays revealed that plakoglobin and p53 associate with the 14-3-3s promoter. Furthermore, luciferase reporter assays showed that p53 transcriptional activity is increased in the presence of plakoglobin. Finally, knockdown of plakoglobin in MCF-7 cells followed by luciferase assays confirmed that p53 transcriptional activity is enhanced in the presence of plakoglobin. Our data suggest that plakoglobin regulates gene expression in conjunction with p53 and that plakoglobin may regulate p53 transcriptional activity, which may account, in part, for the tumor/metastasis suppressor activity of plakoglobin.

Point mutations and gross deletions of PITX2 appear to produce an equivalent haploinsufficiency phenotype. Quantitative PCR is an efficient means of detecting causative PITX2 deletions in patients with AR and may increase the detection rate at this locus.

These analyses demonstrate that hypoxia induces a phenotypic shift, accompanied by modulation of nuclear factor-IB and YY1 activities, in microvascular endothelial cells of the lungs to support VWF promoter activation.