title: Pkd1 and Nek8 mutations affect cell-cell adhesion and cilia in cysts formed in kidney organ cultures.
abstract: Development of novel therapies for polycystic kidney disease (PKD) requires assays that adequately reflect disease biology and are adaptable to high-throughput screening. Here we describe an embryonic cystic kidney organ culture model and demonstrate that a new mutant allele of the Pkd1 gene (Pkd1(tm1Bdgz)) modulates cystogenesis in this model. Cyst formation induced by cAMP is influenced by the dosage of the mutant allele: Pkd1(tm1Bdgz) -/- cultures develop a larger cystic area compared with +/+ counterparts, while Pkd1(tm1Bdgz) +/- cultures show an intermediate phenotype. A similar relationship between the degree of cystogenesis and mutant gene dosage is seen in cystic kidney organ cultures derived from mice with a mutated Nek8 gene (Nek8(jck)). Both Pkd1- and Nek8- cultures display altered cell-cell junctions, with reduced E-cadherin expression and altered desmosomal protein expression, similar to ADPKD epithelia. Additionally, characteristic ciliary abnormalities are identified in cystic kidney cultures, with elevated ciliary polycystin 1 expression in Nek8 homozygous cultures and elevated ciliary Nek8 protein expression in Pkd1 homozygotes. These data suggest that the Nek8 and Pkd1 genes function in a common pathway to regulate cystogenesis. Moreover, compound Pkd1 and Nek8 heterozygous adult mice develop a more aggressive cystic disease than animals with a mutation in either gene alone. Finally, we validate the kidney organ culture cystogenesis assay as a therapeutic testing platform using the CDK inhibitor roscovitine. Therefore, embryonic kidney organ culture represents a relevant model for studying molecular cystogenesis and a rapid tool for the screening for therapies that block cystic growth.
authors.affiliation: Department of Cell Biology, Genzyme Corporation, 5 Mountain Rd., Framingham, MA 01701-9322, USA.
authors.forename: Thomas A
authors.initials: TA
authors.lastname: Natoli
authors.forename: Tiffany C
authors.initials: TC
authors.lastname: Gareski
authors.forename: William R
authors.initials: WR
authors.lastname: Dackowski
authors.forename: Laurie
authors.initials: L
authors.lastname: Smith
authors.forename: Nikolay O
authors.initials: NO
authors.lastname: Bukanov
authors.forename: Ryan J
authors.initials: RJ
authors.lastname: Russo
authors.forename: Hervé
authors.initials: H
authors.lastname: Husson
authors.forename: Douglas
authors.initials: D
authors.lastname: Matthews
authors.forename: Peter
authors.initials: P
authors.lastname: Piepenhagen
authors.forename: Oxana
authors.initials: O
authors.lastname: Ibraghimov-Beskrovnaya
mesh_terms: D000483:Alleles; D000818:Animals; D015820:Cadherins / Q000378:metabolism; D002448:Cell Adhesion / Q000235:genetics / Q000502:physiology*; D002923:Cilia / Q000187:drug effects / Q000378:metabolism*; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors; D003560:Cysts / Q000378:metabolism / Q000503:physiopathology; D004195:Disease Models, Animal; D005260:Female; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D009154:Mutation / Q000235:genetics*; D000072379:NIMA-Related Kinases; D009924:Organ Culture Techniques; D007690:Polycystic Kidney Diseases / Q000378:metabolism* / Q000503:physiopathology; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011494:Protein Kinases / Q000235:genetics* / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D011687:Purines / Q000494:pharmacology; D000077546:Roscovitine; D050396:TRPP Cation Channels / Q000378:metabolism*
chemical_list: D015820:Cadherins; D018844:Cyclin-Dependent Kinases; D000072379:NIMA-Related Kinases; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D011687:Purines; D000077546:Roscovitine; D050396:TRPP Cation Channels; C499219:Nek8 protein, mouse