Autosomal dominant polycystic kidney disease is caused primarily by mutations in two genes, Pkd1 and Pkd2, which function together as a calcium channel complex. Subcellular localization studies suggest that the polycystin2 protein may act in the apical cell membrane associated with cilia or in internal cell membranes as a calcium release channel. However, it is currently not known where polycystin2 channel activity acts in epithelia or which cellular signaling systems may impinge on polycystin2 function. Disruption of zebrafish polycystin2 gene expression using antisense morpholino oligos results in rapid kidney cyst development, randomized organ laterality, hydrocephalus, and body axis curvature. These defects are rescued by co-injection of the human pkd2 mRNA.
In Aim 1. we propose using the zebrafish as an in vivo system for structure function analysis of polycystin2. We hypothesize that specific amino acid motifs in polycystin2 target it to its cellular site of action. By disrupting these either in the endogenous zebrafish polycystin2 or in rescuing human polycystin2 mRNAs we will test whether subcellular localization affects the cellular function of polycystin2.
In Aim 2. we hypothesize that polycystin 2, acting as a calcium channel, mediates the effects of physical forces on the epithelium. We will measure calcium responses in isolated kidney tubules that lack polycystin2 function or that express altered pkd2 alleles. Finally we propose that mutations in polycytin2 interacting proteins or downstream mediators may effect signaling or function of polycystin2.
In Aim 3. we outline a plan to identify cellular components that interact with polycystin2 by screening for dominant suppressors of the zebrafish polycystin2 phenotype. This proposal exploits unique advantages of the zebrafish as a model organism to more fully explore the function of polycystin2 in vivo and further our understanding of the cellular mechanisms of autosomal dominant polycystic kidney disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK070263-02
Application #
6941689
Study Section
Special Emphasis Panel (ZDK1-GRB-6 (J1))
Program Officer
Rasooly, Rebekah S
Project Start
2004-09-01
Project End
2008-08-31
Budget Start
2005-09-01
Budget End
2006-08-31
Support Year
2
Fiscal Year
2005
Total Cost
$258,244
Indirect Cost
Name
Massachusetts General Hospital
Department
Type
DUNS #
073130411
City
Boston
State
MA
Country
United States
Zip Code
02199
Le Corre, Stéphanie; Eyre, David; Drummond, Iain A (2014) Modulation of the secretory pathway rescues zebrafish polycystic kidney disease pathology. J Am Soc Nephrol 25:1749-59
Palmyre, Aurélien; Lee, Jeongeun; Ryklin, Gennadiy et al. (2014) Collective epithelial migration drives kidney repair after acute injury. PLoS One 9:e101304
Slaats, Gisela G; Ghosh, Amiya K; Falke, Lucas L et al. (2014) Nephronophthisis-associated CEP164 regulates cell cycle progression, apoptosis and epithelial-to-mesenchymal transition. PLoS Genet 10:e1004594
Thomas, Sophie; Wright, Kevin J; Le Corre, Stéphanie et al. (2014) A homozygous PDE6D mutation in Joubert syndrome impairs targeting of farnesylated INPP5E protein to the primary cilium. Hum Mutat 35:137-46
Zariwala, Maimoona A; Gee, Heon Yung; Kurkowiak, Małgorzata et al. (2013) ZMYND10 is mutated in primary ciliary dyskinesia and interacts with LRRC6. Am J Hum Genet 93:336-45
Drummond, Iain; Austin-Tse, Christina (2013) Zebrafish cilia. Methods Enzymol 525:219-44
Drummond, Iain A (2012) Cilia functions in development. Curr Opin Cell Biol 24:24-30
Vasilyev, Aleksandr; Liu, Yan; Hellman, Nathan et al. (2012) Mechanical stretch and PI3K signaling link cell migration and proliferation to coordinate epithelial tubule morphogenesis in the zebrafish pronephros. PLoS One 7:e39992
Vasilyev, Aleksandr; Drummond, Iain A (2012) Live imaging kidney development in zebrafish. Methods Mol Biol 886:55-70
Drummond, Iain A (2011) Polycystins, focal adhesions and extracellular matrix interactions. Biochim Biophys Acta 1812:1322-6

Showing the most recent 10 out of 19 publications