CHARACTERIZATION OF A MITOGEN-RESPONSE UNIT IN THE MOUSE LACTOFERRIN GENE PROMOTER

Lactoferrin is present in a variety of tissues and biological fluids; however, the amount differs significantly due to differential expressi ons. We have previously demonstrated that the mouse lactoferrin gene i s regulated by estrogen through an estrogen-response DNA element locat ed at -349, upstream from the transcription start site (+1). In this r eport, we characterized by deletion and mutation analyses a cluster of mitogen-response elements located between -80 and -40 of the mouse la ctoferrin promoter. We demonstrated that the chimeric chloramphenicol acetyltransferase reporter constructs (the -103 to +1 sequence of the mouse lactoferrin gene) containing the mitogen-response unit of the la ctoferrin gene were stimulated by cAMP, forskolin, 12-O-tetradecanoylp horbol-13-acetate, and epidermal growth factor/recombinant transformin g growth factor-alpha (EGF/TGF-alpha) in a time- and dose-dependent ma nner. The sequence at position -52 to -40 (mLF-CRE) of the gene confer red transcriptional activation in the presence of forskolin, cyclic AM P, and 12-O-tetradecanoylphorbol-13-acetate in transiently transfected human endometrium carcinoma RL95-2 cells, whereas the region at -80 t o -60 responded to EGF/TGF-alpha stimulation. Overexpression of the ca talytic unit of protein kinase C or protein kinase A in the RL95-2 cel ls elevated the chloramphenicol acetyltransferase activity of the repo rter construct 5-6-fold. The mobility shift assay suggested that AP1 a nd CREB or related proteins participated in complex formation with the mLF-CRE, whereas different proteins bound to the EGF/TGF-alpha-respon se element.