Biology subjects

Guay-Woodford, L. M.

Publications and source records attributed to Guay-Woodford, L. M..

2 recordsLinked to original sources

bioRxiv2022

Cystin is required for maintaining fibrocystin (FPC) levels and safeguarding proteome integrity in mouse renal epithelial cells Running title: Cystin deficiency reduces FPC levels through enhanced autophagic degradation.

Autosomal recessive polycystic kidney disease (ARPKD) is caused primarily by mutations in PKHD1, encoding fibrocystin (FPC), but Pkhd1 mutant mice fail to express renal cystic disease. In contrast, the renal lesion in Cys1cpk/cpk (cpk) mice with loss of the cystin protein, closely phenocopy ARPKD. Recent identification of patients with CYS1-related ARPKD prompted the investigations described herein. We analyzed cystin and FPC expression in mouse models (cpk, rescued-cpk (r-cpk), Pkhd1 mutants) and cortical collecting duct (CCD) cell lines (wild type (wt), cpk). We found that cystin deficiency led to diminished FPC in both cpk kidneys and CCD cells. In r-cpk kidneys, FPC increased and siRNA of Cys1 in wt CCD cells reduced FPC. Conversely, FPC deficiency in Pkhd1 mutants did not affect cystin levels. Cystin deficiency and the associated reduction in FPC levels impacted the architecture of the primary cilium, but not ciliogenesis. Similar Pkhd1 mRNA levels in wt, cpk kidneys and CCD cells suggested posttranslational mechanisms directed FPC loss and studies of cellular protein degradation systems revealed selective autophagy as a possible mechanism. Loss of FPC from the NEDD4 E3 ubiquitin ligase complexes caused reduced polyubiquitination and elevated levels of functional epithelial sodium channel (NEDD4 target) in cpk cells. We propose that cystin is necessary to stabilize FPC and loss of cystin leads to rapid FPC degradation. FPC removal from E3-ligase complexes alters the cellular proteome and may contribute to cystogenesis through multiple mechanisms, that include MYC transcriptional regulation.

cell biology↗

bioRxiv2020

Cystin gene mutations cause autosomal recessive polycystic kidney disease associated with altered Myc expression

Mutation of the Cys1 gene underlies the renal cystic disease in the Cys1cpk/cpk (cpk) mouse that phenocopies human autosomal recessive polycystic kidney disease (ARPKD). Cystin, the protein product of Cys1, is expressed in the primary apical cilia of renal ductal epithelial cells. In previous studies, we showed that cystin regulates Myc expression via interaction with the tumor suppressor, necdin. Here, we demonstrate rescue of the cpk renal phenotype by kidney-specific expression of a cystin-GFP fusion protein encoded by a transgene integrated into the Rosa26 locus. In addition, we show that expression of the cystin-GFP fusion protein in collecting duct cells down-regulates expression of Myc in cpk kidneys. Finally, we report the first human patient with an ARPKD phenotype due to homozygosity for a predicted deleterious splicing defect in CYS1. These findings suggest that mutations in the Cys1 mouse and CYS1 human orthologues cause an ARPKD phenotype that is driven by overexpression of the Myc proto-oncogene. Translational StatementThe cystin-deficient cpk mouse is a model for the study of autosomal recessive polycystic kidney disease (ARPKD). We show that the cpk mouse phenotype is associated with altered Myc expression. To date, the clinical relevance of cystin deficiency to human disease was unclear, due to the absence of ARPKD cases associated with CYS1 mutations. We report the first case of ARPKD linked to a CYS1 mutation disrupting normal splicing. These findings confirm the relevance of cystin deficiency to human ARPKD, implicate Myc in disease initiation or progression, and validate the cpk mouse as a translationally relevant disease model.

molecular biology↗