title: Expression of podocalyxin inhibits cell-cell adhesion and modifies junctional properties in Madin-Darby canine kidney cells.
abstract: Podocalyxin is a major membrane protein of the glomerular epithelium and is thought to be involved in maintenance of the architecture of the foot processes and filtration slits characteristic of this unique epithelium by virtue of its high negative charge. However, until now there has been no direct evidence for podocalyxin's function. Podocalyxin is a type 1 transmembrane sialoprotein with an N-terminal mucin-like domain. To assess its function, we cloned rat podocalyxin and examined the effects of its expression on the cell adhesion properties of stably transfected Chinese hamster ovary (CHO)-K1 and Madin-Darby canine kidney (MDCK) cells and inducible ecdysone receptor-expressing (EcR)-CHO cells. In a cell aggregation assay, CHO-K1 cells expressing high levels of podocalyxin showed complete inhibition of cell aggregation, and MDCK transfectants showed greatly reduced aggregation ( approximately 60-80%) compared with parental cells. In EcR-CHO cells, the expression level of podocalyxin induced by increasing levels of ecdysone analogue correlated closely with the antiadhesion effect. The inhibitory effect of podocalyxin was reversed by treatment of the cells with Arthrobacter ureafaciens sialidase, indicating that sialic acid is required for inhibition of cell adhesion. Overexpression of podocalyxin also affected transepithelial resistance and the distribution of junctional proteins in MDCK cells by an unknown mechanism that may involve interaction with the actin cytoskeleton. These results provide direct evidence that podocalyxin functions as an antiadhesin that maintains an open filtration pathway between neighboring foot processes in the glomerular epithelium by charge repulsion.
authors.affiliation: Department of Cellular and Molecular Medicine, University of California San Diego, La Jolla, California 92093, USA.
authors.forename: T
authors.initials: T
authors.lastname: Takeda
authors.forename: W Y
authors.initials: WY
authors.lastname: Go
authors.forename: R A
authors.initials: RA
authors.lastname: Orlando
authors.forename: M G
authors.initials: MG
authors.lastname: Farquhar
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001173:Arthrobacter / Q000201:enzymology; D016466:CHO Cells; D002448:Cell Adhesion / Q000502:physiology*; D002449:Cell Aggregation / Q000502:physiology; D002460:Cell Line; D002645:Chickens; D003001:Cloning, Molecular; D006224:Cricetinae; D004285:Dogs; D006031:Glycosylation; D006801:Humans; D007365:Intercellular Junctions / Q000502:physiology*; D007668:Kidney; D008969:Molecular Sequence Data; D009439:Neuraminidase / Q000378:metabolism; D010766:Phosphorylation; D011817:Rabbits; D051381:Rats; D011987:Receptors, Steroid / Q000235:genetics / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D012795:Sialoglycoproteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D014162:Transfection
chemical_list: D011987:Receptors, Steroid; D011993:Recombinant Fusion Proteins; D012795:Sialoglycoproteins; C055469:ecdysone receptor; C001643:podocalyxin; D009439:Neuraminidase