Ischemia and reperfusion enhance ATF-2 and c-Jun binding to cAMP response elements and to an AP-1 binding site from the c-jun promoter.
The transcription factors controlling the complex genetic response to ischemia and their modes of regulation are poorly understood. We found that ATF-2 and c-Jun DNA binding activity is markedly enhanced in post-ischemic kidney or in LLC-PK1 renal tubular epithelial cells exposed to reversible ATP depletion. After 40 min of renal ischemia followed by reperfusion for as little as 5 min, binding of ATF-2 and c-Jun, but not ATF-3 or CREB (cAMP response element binding protein), to oligonucleotides containing either an ATF/cAMP response element (ATF/CRE) or the jun2TRE from the c-jun promoter, was significantly increased. Binding to jun2TRE and ATF/CRE oligonucleotides occurred with an identical time course. In contrast, nuclear protein binding to an oligonucleotide containing a canonical AP-1 element was not detected until 40 min of reperfusion, and although c-Jun was present in the complex, ATF-2 was not. Incubating nuclear extracts from reperfused kidney with protein phosphatase 2A markedly reduced binding to both the ATF/CRE and jun2TRE oligonucleotides, compatible with regulation by an ATF-2 kinase. An ATF-2 kinase, which phosphorylated both the transactivation and DNA binding domains of ATF-2, was activated by reversible ATP depletion. This kinase coeluted on Mono Q column chromatography with a c-Jun amino-terminal kinase and with the peak of stress-activated protein kinase, but not p38, immunoreactivity. In conclusion, DNA binding activity of ATF-2 directed at both ATF/CRE and jun2TRE motifs is modulated in response to the extreme cellular stress of ischemia and reperfusion or reversible ATP depletion. Phosphorylation-dependent activation of the DNA binding activity of ATF-2, which appears to be regulated by the stress-activated protein kinases, may play an important role in the earliest stages of the genetic response to ischemia/reperfusion by targeting ATF-2 and c-Jun to specific promoters, including the c-jun promoter and those containing ATF/CREs.
Medical Services, Massachusetts General Hospital, Charlestown 02129, USA.
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H
Morooka
J V
JV
Bonventre
C M
CM
Pombo
J M
JM
Kyriakis
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T
Force
D051698:Activating Transcription Factor 2; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002852:Chromatography, Ion Exchange; D016384:Consensus Sequence; D017362:Cyclic AMP Response Element-Binding Protein / Q000302:isolation & purification / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism; D004789:Enzyme Activation; D016758:Genes, jun; D007511:Ischemia / Q000378:metabolism*; D007668:Kidney / Q000098:blood supply* / Q000378:metabolism*; D007684:Kidney Tubules / Q000378:metabolism; D016350:Leucine Zippers; D008297:Male; D048056:Mitogen-Activated Protein Kinase 9; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry / Q000378:metabolism; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D011494:Protein Kinases / Q000302:isolation & purification / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000302:isolation & purification / Q000378:metabolism; D016755:Proto-Oncogene Proteins c-jun / Q000302:isolation & purification / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism; D015424:Reperfusion; D018808:Transcription Factor AP-1 / Q000378:metabolism*; D014157:Transcription Factors
D051698:Activating Transcription Factor 2; C497107:Atf2 protein, rat; D017362:Cyclic AMP Response Element-Binding Protein; D004268:DNA-Binding Proteins; D009838:Oligodeoxyribonucleotides; D016755:Proto-Oncogene Proteins c-jun; D011993:Recombinant Fusion Proteins; D018808:Transcription Factor AP-1; D014157:Transcription Factors; D000255:Adenosine Triphosphate; D011494:Protein Kinases; D048056:Mitogen-Activated Protein Kinase 9; D017346:Protein Serine-Threonine Kinases; D020928:Mitogen-Activated Protein Kinases