TGF-beta signaling and its effect on glutaminase expression in LLC-PK1-FBPase+ cells.
During systemic acidosis, renal proximal tubular cells exhibit enhanced rates of bicarbonate and ammonium ion synthesis and undergo extensive hypertrophy. The former adaptations are accomplished, in part, by increased expression of glutaminase (GA). LLC-PK(1)-FBPase+ cells, a gluconeogenic line of porcine kidney cells, exhibit a rapid activation of the ERK1/2 and p38 MAPK pathways and a two- to threefold increase in GA mRNA when transferred to acidic medium (pH 6.9). Transforming growth factor-beta (TGF-beta), a potent activator of MAPK and Smad signaling cascades, also causes extensive renal hypertrophy. Thus the potential role of TGF-beta in the renal response to metabolic acidosis was investigated. Western blot analyses established that in LLC-PK(1)-FBPase+ cells, TGF-beta activated the ERK1/2, p38 MAPK, and Smad1/5/8 pathways, but not the JNK and Smad2/3 pathways. Addition of TGF-beta to cells cultured in normal medium (pH 7.4) produced a steady increase in GA mRNA, resulting in a twofold induction after 18 h. Western blot analysis indicated that treatment with either TGF-beta or acidic medium resulted in an increased level of fibronectin. However, the effects of the two treatments on both GA mRNA and fibronectin levels occurred with different time courses and were additive. In addition, the rates of ammonia production were decreased slightly by addition of TGF-beta. Finally, a GA-luciferase reporter construct, which is activated 3.5-fold by treatment with acidic medium, is not affected by TGF-beta. Therefore, TGF-beta and metabolic acidosis activate some of the same signaling pathways in LLC-PK(1)-FBPase+ cells, but produce separate effects on GA expression.
Andratsch
Manfred
M
Department of Physiology and Medical Physics, Innsbruck Medical University, Fritz-Pregl-Strasse 3, A-6020 Innsbruck, Austria.
Feifel
Elisabeth
E
Taylor
Lynn
L
O'Hayre
Morgan
M
Schramek
Herbert
H
Curthoys
Norman P
NP
Gstraunthaler
Gerhard
G
D000818:Animals; D002460:Cell Line; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005786:Gene Expression Regulation; D005972:Glutaminase / Q000235:genetics* / Q000378:metabolism*; D048670:MAP Kinase Kinase 4 / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000502:physiology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051785:Smad Proteins / Q000378:metabolism; D013552:Swine; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
D012333:RNA, Messenger; D051785:Smad Proteins; D016212:Transforming Growth Factor beta; D048049:Extracellular Signal-Regulated MAP Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D048670:MAP Kinase Kinase 4; D005972:Glutaminase