Cellular and subcellular localization of the ARPKD protein; fibrocystin is expressed on primary cilia

Christopher J. Ward, David Yuan, Tatyana V. Masyuk, Xiaofang Wang, Rachaneekorn Punyashthiti, Shelly Whelan, Robert Bacallao, Roser Torra, Nicholas F. LaRusso, Vicente E. Torres, Peter C. Harris

Research output: Contribution to journalArticle

225 Citations (Scopus)

Abstract

Autosomal recessive polycystic kidney disease (ARPKD) is an infantile form of PKD characterized by fusiform dilation of collecting ducts and congenital hepatic fibrosis. The ARPKD gene, PKHD1, is large (∼470 kb; 67 exons) with a 12 222 bp longest open reading frame, although multiple different splice forms may be generated. The predicted full-length ARPKD protein, fibrocystin, is membrane bound with 4074 amino acids (447 kDa molecular weight). To characterize the pattern of fibrocystin expression we have generated four monoclonal antibodies (mAb) to the cytoplasmic tail of the protein. Western analysis of human kidney membrane protein showed an identical pattern with each mAb; a strongly expressing large product (>450 kDa), consistent with the predicted protein size, and a weaker ∼220 kDa band. The same large product was detected in rat and mouse kidney with lower level expression in liver. To further show that these mAbs recognize fibrocystin, tissue from ARPKD patients was analyzed and no fibrocystin products were detected. Immunohistochemical analysis of the developing kidney showed expression in the branching ureteric bud and collecting ducts, expression that persisted into adulthood. Biliary duct staining was found in the liver, plus staining in the pancreas and developing testis. Immunofluorescence analysis of MDCK cells showed a major site of expression in the primary cilia. Recent studies have associated the disease protein in various human and animal forms of PKD with cilia. The localization of fibrocystin to cilia further strengthens that correlation and indicates that the primary defect in ARPKD may be linked to ciliary dysfunction.

Original languageEnglish
Pages (from-to)2703-2710
Number of pages8
JournalHuman Molecular Genetics
Volume12
Issue number20
DOIs
StatePublished - Oct 15 2003

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Autosomal Recessive Polycystic Kidney
Cilia
Proteins
Kidney
Monoclonal Antibodies
Staining and Labeling
Madin Darby Canine Kidney Cells
Liver
Open Reading Frames
Fluorescent Antibody Technique
Tail
Testis
Dilatation
Pancreas
Exons
Membrane Proteins
Molecular Weight
Amino Acids
Membranes
Genes

ASJC Scopus subject areas

  • Genetics

Cite this

Ward, C. J., Yuan, D., Masyuk, T. V., Wang, X., Punyashthiti, R., Whelan, S., ... Harris, P. C. (2003). Cellular and subcellular localization of the ARPKD protein; fibrocystin is expressed on primary cilia. Human Molecular Genetics, 12(20), 2703-2710. https://doi.org/10.1093/hmg/ddg274

Cellular and subcellular localization of the ARPKD protein; fibrocystin is expressed on primary cilia. / Ward, Christopher J.; Yuan, David; Masyuk, Tatyana V.; Wang, Xiaofang; Punyashthiti, Rachaneekorn; Whelan, Shelly; Bacallao, Robert; Torra, Roser; LaRusso, Nicholas F.; Torres, Vicente E.; Harris, Peter C.

In: Human Molecular Genetics, Vol. 12, No. 20, 15.10.2003, p. 2703-2710.

Research output: Contribution to journalArticle

Ward, CJ, Yuan, D, Masyuk, TV, Wang, X, Punyashthiti, R, Whelan, S, Bacallao, R, Torra, R, LaRusso, NF, Torres, VE & Harris, PC 2003, 'Cellular and subcellular localization of the ARPKD protein; fibrocystin is expressed on primary cilia', Human Molecular Genetics, vol. 12, no. 20, pp. 2703-2710. https://doi.org/10.1093/hmg/ddg274
Ward, Christopher J. ; Yuan, David ; Masyuk, Tatyana V. ; Wang, Xiaofang ; Punyashthiti, Rachaneekorn ; Whelan, Shelly ; Bacallao, Robert ; Torra, Roser ; LaRusso, Nicholas F. ; Torres, Vicente E. ; Harris, Peter C. / Cellular and subcellular localization of the ARPKD protein; fibrocystin is expressed on primary cilia. In: Human Molecular Genetics. 2003 ; Vol. 12, No. 20. pp. 2703-2710.
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AU - Punyashthiti, Rachaneekorn

AU - Whelan, Shelly

AU - Bacallao, Robert

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