Expression and function of Ets-1 during experimental acute renal failure in rats.
The Ets family of transcription factors is defined by a conserved DNA-binding Ets domain that forms a winged helix-turn-helix structure motif. The Ets family is involved in a diverse array of biologic functions, including cellular growth, migration, and differentiation. The hypothesis in this study was that Ets-1 is re-expressed during regeneration after acute renal failure (ARF) and plays a key role in the transcriptional regulation of cyclin D1 and the cell cycle progression in renal tubular cells. For clarifying the significance of Ets-1 in ARF, a rat ARF model in vivo and LLC-PK1 cells as an in vitro model were used. After the left rat renal artery was clamped for 1 h, the whole kidney homogenate was examined and total RNA was extracted at 6, 12, 24, 48, and 72 h after reperfusion by Western blot analysis and real-time reverse transcription-PCR. Ets-1 mRNA and protein expression were strongly increased at 6 to 24 h after the ischemia, respectively. The expression of hypoxia-inducible factor-1alpha was increased dramatically as early as 6 h after ischemia-reperfusion and decreased at 48 and 72 h after ischemia-reperfusion. In the immunohistologic examination, Ets-1 was expressed in the proximal tubules and coexpressed with proliferating cell nuclear antigen (PCNA). Furthermore, overexpression of Ets-1 promoted the cell cycle and increased the promoter activity and protein expression of cyclin D1 in LLC-PK1 cells. Ets-1 promoter activity increased between 3 and 6 h in hypoxia, and hypoxia also induced changes in the Ets-1 protein level in LLC-PK1 cells. The Ets-1 induction by hypoxia was abolished by the transfection of dominant-negative hypoxia-inducible factor-1alpha. A gel shift assay demonstrated that Ets-1 binds to the ets-1 binding site of the cyclin D1 promoter in the ischemia-reperfusion condition. Overexpression of Ets-1 did not significantly change the caspase 3 activity or the value of cell death ELISA in LLC-PK1 cells. Taken together, these data suggest that Ets-1 plays a key role in the cell-cycle progression of renal tubules in ARF. The Ets-1 pathway may regulate the transcription of cyclin D1 and control the regeneration of renal tubules in ARF.
Tanaka
Hiroyuki
H
Homeostasis Medicine and Nephrology, Tokyo Medical and Dental University, 5-45 Yushima 1-chome, Bunkyo-ku, Tokyo 113-8519, Japan.
Terada
Yoshio
Y
Kobayashi
Takahiko
T
Okado
Tomokazu
T
Inoshita
Seiji
S
Kuwahara
Michio
M
Seth
Arun
A
Sato
Yasufumi
Y
Sasaki
Sei
S
D058186:Acute Kidney Injury / Q000503:physiopathology*; D000818:Animals; D017209:Apoptosis / Q000502:physiology; D015153:Blotting, Western; D002455:Cell Division / Q000502:physiology; D019938:Cyclin D1 / Q000235:genetics; D024202:Electrophoretic Mobility Shift Assay; D015870:Gene Expression; D000860:Hypoxia / Q000503:physiopathology; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007150:Immunohistochemistry; D018374:LLC-PK1 Cells; D008297:Male; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic / Q000502:physiology; D050784:Proto-Oncogene Protein c-ets-1; D011518:Proto-Oncogene Proteins / Q000235:genetics* / Q000378:metabolism*; D050783:Proto-Oncogene Proteins c-ets; D051381:Rats; D017207:Rats, Sprague-Dawley; D012038:Regeneration / Q000502:physiology; D013552:Swine; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism*; D014158:Transcription, Genetic / Q000502:physiology
C494043:Ets1 protein, rat; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D050784:Proto-Oncogene Protein c-ets-1; D011518:Proto-Oncogene Proteins; D050783:Proto-Oncogene Proteins c-ets; D014157:Transcription Factors; D019938:Cyclin D1