Regulation of the vav proto-oncogene by LKLF

Diane J. Denkinger, Allison M Cushman-Vokoun, Rodney S. Kawahara

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

The transcriptional induction of the vav proto-oncogene coincides with the first appearance of the definitive hematopoietic stem cell in the aorta-gonad-mesonephros region. Vav promoter activity was dependent on a previously identified 23 bp DNA segment containing PU.1 and Runx1/AML-1 binding sites and on a newly identified, highly conserved, 12 bp DNA segment (Box B). The sequence of Box B was identical in the human, mouse and rat species. Mutation of the CACCC core sequence led to diminished vav promoter activity. A protein complex which bound to Box B was found in hematopoietic cells but not in cells which did not express vav. A double-stranded oligonucleotide containing a mutation of the CACCC core was less effective in electro-mobility shift assay competitions than the wild-type sequence. UV crosslinking studies identified a 37 kDa DNA binding protein which interacted with Box B in U937 cells. Antibody supershift assays identified this protein as lung Krüppel-like factor (LKLF). LKLF, expressed as a glutathione S-transferase fusion protein, was capable of binding to Box B. A dominant-negative LKLF was able to inhibit the expression of enhanced green fluorescent protein by the vav promoter and chromatin immunoprecipitations detected LKLF bound to the vav promoter in U937 but not HeLa cells. These in vitro results suggest future in vivo experiments to examine the role of LKLF, a gene required for vasculogenesis, in the induction of vav during the genesis of the definitive hematopoietic stem cell from the vascular endothelium.

Original languageEnglish (US)
Pages (from-to)133-142
Number of pages10
JournalGene
Volume281
Issue number1-2
DOIs
StatePublished - Dec 27 2001

Fingerprint

Proto-Oncogenes
Lung
Hematopoietic Stem Cells
Mesonephros
U937 Cells
Mutation
Proteins
Chromatin Immunoprecipitation
DNA
Vascular Endothelium
DNA-Binding Proteins
Gonads
Electrophoretic Mobility Shift Assay
Glutathione Transferase
HeLa Cells
Oligonucleotides
Aorta
Binding Sites
Antibodies
Genes

Keywords

  • Gene expression
  • Hematopoiesis
  • Stem cells
  • Transcription

ASJC Scopus subject areas

  • Genetics

Cite this

Regulation of the vav proto-oncogene by LKLF. / Denkinger, Diane J.; Cushman-Vokoun, Allison M; Kawahara, Rodney S.

In: Gene, Vol. 281, No. 1-2, 27.12.2001, p. 133-142.

Research output: Contribution to journalArticle

Denkinger, Diane J. ; Cushman-Vokoun, Allison M ; Kawahara, Rodney S. / Regulation of the vav proto-oncogene by LKLF. In: Gene. 2001 ; Vol. 281, No. 1-2. pp. 133-142.
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