SREBP-1 activation by glucose mediates TGF-β upregulation in mesangial cells.
Glomerular matrix accumulation is a hallmark of diabetic nephropathy. Recent studies showed that overexpression of the transcription factor sterol-responsive element-binding protein (SREBP)-1 induces pathology reminiscent of diabetic nephropathy, and SREBP-1 upregulation was observed in diabetic kidneys. We thus studied whether SREBP-1 is activated by high glucose (HG) and mediates its profibrogenic responses. In primary rat mesangial cells, HG activated SREBP-1 by 30 min, seen by the appearance of its cleaved nuclear form (nSREBP-1), EMSA, and by activation of an SREBP-1 response element (SRE)-driven green fluorescent protein construct. Activation was dose dependent and not induced by an osmotic control. Site 1 protease was required, since its inhibition by AEBSF prevented SREBP-1 activation. SCAP, the ER-associated chaperone for SREBP-1, was also necessary since its inhibitor fatostatin also blocked SREBP-1 activation. Signaling through the EGFR/phosphatidylinositol 3-kinase (PI3K) pathway, which we previously showed mediates HG-induced TGF-β1 upregulation, and through RhoA, were upstream of SREBP-1 activation (Wu D, Peng F, Zhang B, Ingram AJ, Gao B, Krepinsky JC. Diabetologia 50: 2008-2018, 2007; Wu D, Peng F, Zhang B, Ingram AJ, Kelly DJ, Gilbert RE, Gao B, Krepinsky JC. J Am Soc Nephrol 20: 554-566, 2009). Fatostatin and AEBSF prevented HG-induced TGF-β1 upregulation by Northern blot analysis, and HG-induced TGF-β1 promoter activation was inhibited by both fatostatin and dominant negative SREBP-1a. Chromatin immunoprecipitation analysis confirmed that HG led to SREBP-1 binding to the TGF-β1 promoter in a region containing a putative SREBP-1 binding site (SRE). Thus HG-induced SREBP-1 activation requires EGFR/PI3K/RhoA signaling and SCAP-mediated transport to the Golgi for its proteolytic cleavage. Activated SREBP-1 binds to the TGF-β promoter, resulting in TGF-β1 upregulation in response to HG. SREBP-1 thus provides a potential novel therapeutic target for the treatment of diabetic nephropathy.
St. Joseph's Hospital, Hamilton, ON, Canada.
Lalita
L
Uttarwar
Bo
B
Gao
Alistair J
AJ
Ingram
Joan C
JC
Krepinsky
D000818:Animals; D003921:Diabetes Mellitus, Experimental / Q000378:metabolism; D003928:Diabetic Nephropathies / Q000378:metabolism*; D066246:ErbB Receptors / Q000378:metabolism; D005947:Glucose / Q000378:metabolism* / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D050527:Mesangial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D051379:Mice; D008815:Mice, Inbred Strains; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D061251:Primary Cell Culture; D011401:Promoter Regions, Genetic / Q000502:physiology; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D051780:Sterol Regulatory Element Binding Protein 1 / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation / Q000187:drug effects / Q000502:physiology; D053773:Transforming Growth Factor beta1 / Q000235:genetics / Q000378:metabolism*; D015854:Up-Regulation / Q000187:drug effects / Q000502:physiology; D020742:rhoA GTP-Binding Protein / Q000378:metabolism
D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; C102196:SREBP cleavage-activating protein; C497388:Srebf1 protein, mouse; C497389:Srebf1 protein, rat; D051780:Sterol Regulatory Element Binding Protein 1; D053773:Transforming Growth Factor beta1; D019869:Phosphatidylinositol 3-Kinases; D066246:ErbB Receptors; D051057:Proto-Oncogene Proteins c-akt; D020742:rhoA GTP-Binding Protein; D005947:Glucose