Cloning and functional analysis of the hematopoietic cell-specific phospholipase Cγ2promoter

Abstract

Phospholipase Cγ ² (PLCγ ²) is a phospholipid-converting enzyme which, upon receptor stimulation, is activated within membrane-bound signalling complexes. In contrast to the highly ubiquitous PLCγ ¹, PLCγ ² is expressed predominantly in B-lymphocytes. Associated with antigen-coupling receptors it is activated by tyrosine phosphorylation after the triggering of B-cell surface immunoglobulin. We have cloned and sequenced the human PLCγ ² promoter. Primer extension analysis reveals the existence of a major transcriptional start site. The TATA-less promoter contains G+C-rich stretches with a cluster of contiguous SP1 consensus sites, an NF1, and an AP2 site between by −220 to −70. A construct containing the region from −189 to +78 confers full promoter activity, as shown by fusion to a luciferase reporter gene construct. The distal part of the promoter between by −662 to −293 containing an SRE, EBF and CACCC box contributed negatively to promoter activity in the B-cell line Raji but not in three adherent cell lines. In Raji cells, PLCγ ² mRNA is expressed at low levels with a half life greater than 4 h. After treatment with serum, TPA, retinoic acid, or with 5-azacytidine increased levels of PLCγ ² mRNA were induced in B-cells.