Regulation of type II transforming-growth-factor-beta receptors by protein kinase C iota.
TGF-beta (transforming growth factor-beta) is implicated in the pathogenesis of diabetic nephropathy. We previously demonstrated that up-regulation of type II TGF-beta receptor (TbetaRII) induced by high glucose might contribute to distal tubular hypertrophy [Yang, Guh, Yang, Lai, Tsai, Hung, Chang and Chuang (1998) J. Am. Soc. Nephrol. 9, 182-193]. We have elucidated the mechanism by using cultured Madin-Darby canine kidney cells. Enhancer assay and electrophoretic-mobility-shift assay were used to estimate the involvement of transcription factors. Western blotting and an in vitro kinase assay were used to evaluate the level and activity of protein kinase. We showed that glucose (100-900 mg/dl) induced an increase in mRNA level and promoter activity of TbetaRII (note: 'mg/dl' are the units commonly used in diabetes studies). The promoter region -209 to -177 appeared to contribute to positive transactivation of TbetaRII promoter by comparing five TbetaRII-promoter-CAT (chloramphenicol acetyl-transferase) plasmids. Moreover, the transcription factor AP-1 (activator protein 1) was significantly activated and specifically binds to TbetaRII promoter (-209 to -177). More importantly, we found that atypical PKC iota might be pivotal for high glucose-induced increase in both AP-1 binding and TbetaRII promoter activity. First, high glucose induced cytosolic translocation, activation and autophosphorylation of PKC iota. Secondly, antisense PKC iota expression plasmids attenuated high-glucose-induced increase in AP-1 binding and TbetaRII promoter activity; moreover, sense PKC iota expression plasmids enhanced these instead. Finally, we showed that antisense PKC iota expression plasmids might partly attenuate a high-glucose/TGF-beta1-induced increase in fibronectin. We conclude that PKC iota might mediate high-glucose-induced increase in TbetaRII promoter activity. In addition, antisense PKC iota expression plasmid effectively suppressed up-regulation of TbetaRII and fibronectin in hyperglycaemic distal-tubule cells.
Chuang
Lea-Yea
LY
Department of Biochemistry, Kaohsiung Medical University, Tainan, Taiwan.
Guh
Jinn-Yuh
JY
Liu
Shu-Fen
SF
Hung
Min-Yuan
MY
Liao
Tung-Nan
TN
Chiang
Tai-An
TA
Huang
Jau-Shyang
JS
Huang
Yu-Lun
YL
Lin
Chi-Fong
CF
Yang
Yu-Lin
YL
D000704:Analysis of Variance; D000818:Animals; D015152:Blotting, Northern; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D003600:Cytosol / Q000201:enzymology; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D005786:Gene Expression Regulation / Q000187:drug effects; D005947:Glucose / Q000494:pharmacology; D015151:Immunoblotting; D007527:Isoenzymes / Q000378:metabolism*; D007686:Kidney Tubules, Distal / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding / Q000187:drug effects; D011493:Protein Kinase C / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D021381:Protein Transport / Q000187:drug effects; D012333:RNA, Messenger / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D000077294:Receptor, Transforming Growth Factor-beta Type II; D018125:Receptors, Transforming Growth Factor beta / Q000235:genetics* / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D018808:Transcription Factor AP-1 / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism; D000099293:Protein Kinase C-lambda
D015500:Chloramphenicol O-Acetyltransferase; D005947:Glucose; D007527:Isoenzymes; D011493:Protein Kinase C; D017346:Protein Serine-Threonine Kinases; D012333:RNA, Messenger; D000077294:Receptor, Transforming Growth Factor-beta Type II; D018125:Receptors, Transforming Growth Factor beta; D011993:Recombinant Fusion Proteins; D018808:Transcription Factor AP-1; D014157:Transcription Factors; D000099293:Protein Kinase C-lambda