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Am J Physiol Renal Physiol 280: F980-F988, 2001;
0363-6127/01 $5.00
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Vol. 280, Issue 6, F980-F988, June 2001

Angiotensin IV induces tyrosine phosphorylation of focal adhesion kinase and paxillin in proximal tubule cells

Jian-Kang Chen1, Joe Zimpelmann1, Raymond C. Harris2, and Kevin D. Burns1

1 Division of Nephrology, Department of Medicine, Kidney Research Center, Ottawa Hospital Research Institute, University of Ottawa, Ottawa, Ontario, Canada K1H 8L6; and 2 Department of Medicine, Vanderbilt University, Nashville, Tennessee 37232

Angiotensin IV (ANG IV), the COOH-terminal hexapeptide fragment of angiotensin II (ANG II), binds to specific sites in the kidney, distinct from type 1 (AT1) and type 2 (AT2) receptors and designated type 4 (AT4) receptors. We determined signaling pathways for ANG IV in a proximal tubular cell line, LLC-PK1/Cl4. In these cells, we found no specific binding of [125I]-ANG II. In contrast, ANG IV dose dependently competed for [125I]-labeled ANG IV binding, with no displacement by either ANG II, the AT1 receptor antagonist losartan, or the AT2 antagonist PD-123319. Saturation binding indicated the presence of AT4 receptors of high affinity [dissociation constant (Kd) = 1.4 nM]. ANG IV did not affect cAMP or cGMP production and did not increase cytosolic calcium concentration in these cells. In contrast, immunoprecipitation and immunoblotting studies revealed that ANG IV caused dose-dependent tyrosine phosphorylation of p125-focal adhesion kinase (p125-FAK) and p68-paxillin within 2 min, with maximal stimulation at 30 min. ANG IV-stimulated tyrosine phosphorylation of p125-FAK and paxillin was not affected by pretreatment with either losartan or PD-123319, and ANG II (10-7 M) did not induce protein tyrosine phosphorylation. Our results indicate that LLC-PK1/Cl4 cells express ANG IV receptors, which we demonstrate for the first time are linked to tyrosine phosphorylation of focal adhesion-associated proteins. This suggests that ANG IV, a product of ANG II metabolism, may regulate function of the focal adhesion complex in proximal tubule cells.

renin-angiotensin system; receptor; signaling; tyrosine kinase; renal proximal tubular epithelial cells


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