Transcriptional activation of human pregnancy-specific glycoprotein 5 gene by SP1 through binding to a GGGCGTG motif

NORES, R.; PATRITO, L.; PANZETTA-DUTARI, G.

Córdoba

Jornada; Primeras Jornadas de Posgrado de la Facultad de Ciencias Químicas; 2003

Departamento de Posgrado de la Facultad de Ciencias Químicas (UNC)

The human pregnancy-specific glycoprotein 5 gene (PSG5) expression is maximal in placenta and is regulated mainly during transcription. Its minimal promoter lacks TATA-box and large GC-rich sequences, and is active in both PSG-producing and non-producing cell lines. Footprinting assays within the 5’ proximal regulatory region revealed the presence of four cis-acting elements. Now, we have investigated the molecular factors that recognize FP1 element. The incubation of protein extracts of different cell types with FP1 probe yielded two specific DNA-protein complexes, A and B, that were abolished by an excess of unlabelled FP1 competitor DNA but not by mutated FP1 or unrelated oligonucleotides. Complexes were absent when extracts of Drosophila Schneider cells, devoid of Sp1 factor, where employed. In addition, the major complex (A) was competed by a consensus Sp1-binding oligonucleotide and its intensity was increased in EMSA performed with cells transfected with a Sp1 expression plasmid. South-western blots and UV-crosslinking assays revealed that a protein of 104 ± 5 KDa is involved in complexes formation. This molecular weight is in good agreement with the molecular weight reported for Sp1 (106 KDa). Transient-expression assays of PSG5-CAT constructs cotransfected with Sp1 in COS7 cells showed increased transcriptional activity. These results establish that Sp1 binds to the regulatory region and positively regulates the expression of the PSG5 gene.