Nedd4-2 functionally interacts with ClC-5: involvement in constitutive albumin endocytosis in proximal tubule cells

Hryciw, D, Ekberg, J, Lee, A, Lensink, I, Kumar, S, Guggino, W, Cook, D, Pollock, C and Poronnik, P 2004, 'Nedd4-2 functionally interacts with ClC-5: involvement in constitutive albumin endocytosis in proximal tubule cells', Journal of Biological Chemistry, vol. 279, no. 53, pp. 54996-55007.


Document type: Journal Article
Collection: Journal Articles

Title Nedd4-2 functionally interacts with ClC-5: involvement in constitutive albumin endocytosis in proximal tubule cells
Author(s) Hryciw, D
Ekberg, J
Lee, A
Lensink, I
Kumar, S
Guggino, W
Cook, D
Pollock, C
Poronnik, P
Year 2004
Journal name Journal of Biological Chemistry
Volume number 279
Issue number 53
Start page 54996
End page 55007
Total pages 12
Publisher The American Society for Biochemistry and Molecular Biology
Abstract Constitutive albumin uptake by the proximal tubule is achieved by a receptor-mediated process in which the Cl- channel, ClC-5, plays an obligate role. Here we investigated the functional interaction between ClC-5 and ubiquitin ligases Nedd4 and Nedd4-2 and their role in albumin uptake in opossum kidney proximal tubule (OK) cells. In vivo immunoprecipitation using an anti-HECT antibody demonstrated that ClC-5 bound to ubiquitin ligases, whereas glutathione S-transferase pull-downs confirmed that the C terminus of ClC-5 bound both Nedd4 and Nedd4-2. Nedd4-2 alone was able to alter ClC-5 currents in Xenopus oocytes by decreasing cell surface expression of ClC-5. In OK cells, a physiological concentration of albumin (10 ?g/ml) rapidly increased cell surface expression of ClC-5, which was also accompanied by the ubiquitination of ClC-5. Albumin uptake was reduced by inhibiting either the lysosome or proteasome. Total levels of Nedd4-2 and proteasome activity also increased rapidly in response to albumin. Overexpression of ligase defective Nedd4-2 or knockdown of endogenous Nedd4-2 with small interfering RNA resulted in significant decreases in albumin uptake. In contrast, pathophysiological concentrations of albumin (100 and 1000 ?g/ml) reduced the levels of ClC-5 and Nedd4-2 and the activity of the proteasome to the levels seen in the absence of albumin. These data demonstrate that normal constitutive uptake of albumin by the proximal tubule requires Nedd4-2, which may act via ubiquitination to shunt ClC-5 into the endocytic pathway.
Subject Chemical Engineering not elsewhere classified
Keyword(s) animal cell
article
carboxy terminal sequence
cell surface
chloride channel 5
chloride current
controlled study
endocytosis
gene
gene overexpression
immunoprecipitation
in vivo study
kidney proximal tubule
lysosome
molecular mechanics
nedd 4 2 gene
nonhuman
oocyte
opossum
pathophysiology
priority journal
protein expression
protein interaction
protein protein interaction
protein transport
ubiquitination
Xenopus
ISSN 0021-9258
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