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Volume 271, Number 47, Issue of November 22, 1996 pp. 29688-29697
©1996 by The American Society for Biochemistry and Molecular Biology, Inc.

Expression of a 74-kDa Nuclear Factor 1 (NF1) Protein Is Induced in Mouse Mammary Gland Involution
INVOLUTION-ENHANCED OCCUPATION OF A TWIN NF1 BINDING ELEMENT IN THE TESTOSTERONE-REPRESSED PROSTATE MESSAGE-2/CLUSTERIN PROMOTER

(Received for publication, May 30, 1996, and in revised form, August 28, 1996)

Eileen E. M. Furlong Dagger , Niamh K. Keon Dagger , Fiona D. Thornton Dagger , Theo Rein § and Finian Martin Dagger

From the Dagger  Department of Pharmacology and Biotechnology Center, University College Dublin, Dublin 4, Ireland and § Institute of Biochemistry, Ludwig-Maximilians University, D-8000 Munich 2, Germany

Testosterone repressed prostate message-2 (TRPM-2)/clusterin gene expression is rapidly induced in early involution of the mouse mammary gland, after weaning, and in the rat ventral prostate, after castration. A search for involution-enhanced DNaseI footprints in the proximal mouse TRPM-2/clusterin gene promoter led to the identification and characterization (by DNase I footprinting and EMSA) of a twin nuclear factor 1 (NF1) binding element at -356/-309, relative to the proposed transcription start site; nuclear extracts from 2-day involuting mouse mammary gland showed an enhanced footprint over the proximal NF1 element; extracts from involuting prostate showed enhanced occupancy of both NF1 binding elements. Subsequent EMSA and Western analysis led to the detection of a 74-kDa NF1 protein whose expression is triggered in early involution in the mouse mammary gland; such an induced protein is not found in the involuting rat ventral prostate. This protein was not found in lactation where three other NF1 proteins of 114, 68, and 46 kDa were detected. Reiteration of the epithelial cell apoptosis associated with early mammary gland involution, in vitro, in a primary cell culture system, triggered the appearance of the 74-kDa NF1. Overlaying the cells with laminin-rich extracellular matrix suppressed the apoptosis and the expression of the 74-kDa NF1 and, in the presence of lactogenic hormones, initiated milk protein gene expression and the expression of two of the lactation-associated NF1 proteins (68 and 46 kDa). This study, thus, identifies for the first time the occurrence of a switch in expression of different members of the family of NF1 transcription factors as mammary epithelial cells move from the differentiated to the involution/apoptotic state, and it is likely that the involution-specific 74-kDa NF1 accounts for the enhanced NF1 footprint detected on the TRPM-2/clusterin promoter with extracts of mouse mammary gland.


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