Journal
BIOCHEMICAL JOURNAL
Volume 387, Issue -, Pages 239-246Publisher
PORTLAND PRESS LTD
DOI: 10.1042/BJ20041458
Keywords
cytosolic phospholipase A(2) (cPLA(2)); gene transcription; lung Kruppel-like factor (LKLF); non-small-cell lung cancer (NSCLC); small interfering RNA (siRNA); yeast one-hybrid
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Funding
- NCI NIH HHS [P50 CA058187, CA58187, CA108610, R01 CA108610, CA103618, R01 CA103618] Funding Source: Medline
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Increased expression of cPLA, (cytosolic phospholipase A(2)) has been shown to be the cause of tumorigenesis of NSCLC (non-small-cell lung cancer). Our laboratory has previously demonstrated that oncogenic forms of Ras increase transcription of cPLA(2) in normal lung epithelial cells and NSCLC lines through activation of the ERK (extracellular-signal-regulated kinase) and JNK (c-Jun N-terminal kinase) MAPK (mitogen-activated protein kinase) family. We have also defined a minimal region of the cPLA(2) promoter that is critical for this induction. To identify potential transcription factors that bind to this region and regulate expression, a yeast one-hybrid screen was performed with a rat lung cDNA library. Multiple members of the Kruppel family were identified, with LKLF (lung Kruppel-like factor) being isolated a number of times. Overexpression of LKLF in lung epithelial cells or Drosophila SL-2 cells increased cPLA, promoter activity. Conversely, expression of a dominant negative form of LKLF inhibited induction of cPLA promoter activity by oncogenic Ras in normal lung epithelial cells and NSCLC. By electrophoretic mobility-shift assay analysis, it was found that LKLF bound to a GC-rich region of the cPLA2 promoter located between - 37 and - 30 upstream from the transcription start site. Expression of siRNA (small interfering RNA) directed against LKLF inhibited basal expression of cPLA, in lung epithelial cells and blocked induction by H-Ras. In NSCLC, siRNA against LKLF co-operated with siRNA against Sp1 (stimulatory protein 1) to inhibit cPLA, promoter activity. Finally, recombinant LKLF was a substrate for ERKs. These results indicate that LKLF is an important regulator of cPLA, expression and participates in the induction of this protein, which is critical for increased eicosanoid production associated with lung tumorigenesis.
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