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Cloning of the human activated leukocyte cell adhesion molecule promoter and identification of its tissue-independent transcriptional activation by Sp1

Abstract

Activated leukocyte cell adhesion molecule (ALCAM) belongs to the immunoglobulin cell adhesion molecule super family. ALCAM is implicated in tumor progression, inflammation, and the differentiation of hematopoietic stem cells. Hitherto, the identity of regulatory DNA elements and cognate transcription factors responsible for ALCAM gene expression remained unknown. In this report, the human ALCAM promoter was cloned and its transcriptional mechanisms elucidated. The promoter is TATA-less and contains multiple GC-boxes. A proximal 650-bp promoter fragment conferred tissue-independent activation, whereas two contiguous regions upstream of this region negatively influenced promoter activity in a tissue-specific manner. The positive regulatory promoter region was mapped to a core 50 base pair sequence containing a conical Sp1 element. Mutation analysis revealed that this element alone or in tandem with elements immediately upstream was required for maximal promoter activity. Chromatin analysis revealed that Sp1 binds exclusively to the canonical binding sequence in vivo, but not to DNA sequence immediately upstream. Finally, we showed that over-expression of Sp1 significantly increased the basal promoter activity. Thus, Sp1 activated the ALCAM promoter in most cells. These findings have important ramifications for unraveling the roles of ALCAM in inflammation and tumorigenesis.

Abbreviations

ALCAM:

activated leukocyte cell adhesion molecule

CHIP:

chromatin immunoprecipitation

EMSA:

electrophoretic mobility shift assay

PAECs:

pulmon. artery endothelial cells

PMVECs:

pulmon. micro-vascular endothelial cells

pN3Sp1:

plasmid DNA expressing Sp1

Sp1:

specificity protein 1

TBP:

TATA-box binding protein

TFIIB:

transcription factor IIB

TRANSFAC:

TRANScription FACtor database

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Correspondence to Solomon F. Ofori-Acquah.

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Tan, F., Mbunkui, F. & Ofori-Acquah, S.F. Cloning of the human activated leukocyte cell adhesion molecule promoter and identification of its tissue-independent transcriptional activation by Sp1. Cell Mol Biol Lett 17, 571–585 (2012). https://doi.org/10.2478/s11658-012-0028-1

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  • DOI: https://doi.org/10.2478/s11658-012-0028-1

Key words

  • ALCAM
  • Cis elements
  • Endothelial cells
  • Epithelial cells
  • Hematopoietic cells
  • Promoter activity
  • Sp1
  • Transcriptional regulation