TY - JOUR
T1 - Profiling of the renal kinome
T2 - A novel tool to identify protein kinases involved in angiotensin II-dependent hypertensive renal damage
AU - De Borst, Martin H.
AU - Diks, Sander H.
AU - Bolbrinker, Juliane
AU - Schellings, Mark W.
AU - Van Dalen, Mario B.A.
AU - Peppelenbosch, Maikel P.
AU - Kreutz, Reinhold
AU - Pinto, Yigal M.
AU - Navis, Gerjan
AU - Van Goor, Harry
PY - 2007/7
Y1 - 2007/7
N2 - Regulation of protein kinase activities is crucial in both physiology and disease, but analysis is hampered by the multitude and complexity of kinase networks. We used novel peptide array chips containing 1,152 known kinase substrate sequences to profile different kinase activities in renal lysates from homozygous Ren2 rats, a model characterized by hypertension and angiotensin II (ANG II)-mediated renal fibrosis, compared with Sprague-Dawley (SD) control rats and Ren2 rats treated with an angiotensin-converting enzyme inhibitor (ACEi). Five-wk-old homozygous Ren2 rats were left untreated or treated with the ACEi ramipril (1 mg·kg-1·day-1) for 4 wk; age-matched SD rats served as controls (n = 5 each). Peptide array chips were incubated with renal cortical lysates in the presence of radioactively labeled ATP. Radioactivity incorporated into the substrate motifs was measured to quantify kinase activity. A number of kinases with modulated activities, which might contribute to renal damage, were validated by Western blotting, immunoprecipitation, and immunohistochemistry. Relevant kinases identified by the peptide array and confirmed using conventional techniques included p38 MAP kinase and PDGF receptor-β, which were increased in Ren2 and reversed by ACEi. Furthermore, insulin receptor signaling was reduced in Ren2 compared with control rats, and G protein-coupled receptor kinase (GRK) activity decreased in Ren2 + ACEi compared with untreated Ren2 rats. Array-based profiling of tissue kinase activities in ANG II-mediated renal damage provides a powerful tool for identification of relevant kinase pathways in vivo and may lead to novel strategies for therapy.
AB - Regulation of protein kinase activities is crucial in both physiology and disease, but analysis is hampered by the multitude and complexity of kinase networks. We used novel peptide array chips containing 1,152 known kinase substrate sequences to profile different kinase activities in renal lysates from homozygous Ren2 rats, a model characterized by hypertension and angiotensin II (ANG II)-mediated renal fibrosis, compared with Sprague-Dawley (SD) control rats and Ren2 rats treated with an angiotensin-converting enzyme inhibitor (ACEi). Five-wk-old homozygous Ren2 rats were left untreated or treated with the ACEi ramipril (1 mg·kg-1·day-1) for 4 wk; age-matched SD rats served as controls (n = 5 each). Peptide array chips were incubated with renal cortical lysates in the presence of radioactively labeled ATP. Radioactivity incorporated into the substrate motifs was measured to quantify kinase activity. A number of kinases with modulated activities, which might contribute to renal damage, were validated by Western blotting, immunoprecipitation, and immunohistochemistry. Relevant kinases identified by the peptide array and confirmed using conventional techniques included p38 MAP kinase and PDGF receptor-β, which were increased in Ren2 and reversed by ACEi. Furthermore, insulin receptor signaling was reduced in Ren2 compared with control rats, and G protein-coupled receptor kinase (GRK) activity decreased in Ren2 + ACEi compared with untreated Ren2 rats. Array-based profiling of tissue kinase activities in ANG II-mediated renal damage provides a powerful tool for identification of relevant kinase pathways in vivo and may lead to novel strategies for therapy.
UR - http://www.scopus.com/inward/record.url?scp=34548039258&partnerID=8YFLogxK
U2 - 10.1152/ajprenal.00367.2006
DO - 10.1152/ajprenal.00367.2006
M3 - Article
C2 - 17429032
AN - SCOPUS:34548039258
SN - 1931-857X
VL - 293
SP - F428-F437
JO - American Journal of Physiology - Renal Physiology
JF - American Journal of Physiology - Renal Physiology
IS - 1
ER -