The hamster ductus deferns smooth tumor cell line (DDT-1-MF-2) contains androgen receptors and is stimulated by androgens to grow. In the presence of the androgen methyltrienolene, the level of androgen receptor increases to a steady state concentration of 20 fmoles/Mug DNA (2.0 mg of receptor/10-11 cells). The androgen receptor from these cells will be purified by a combination of preparative denaturing Slab gel electrophoresis, affinity chromatography, photoaffinity labelling, and preparative two-dimensional polyacrylamide gel electrophoresis. A phage Lambdagt 11 complementary DNA library with over 250,000 independent recombinant has been prepared from messenger RNA, isolated from androgen stimulated DDT-1 cells. This library has been shown to contain full length inserts to other low abundance messenger RNAs. The pure androgen receptor will be used to determine amino acid composition and the amino acid sequence of various peptides. Several oligonucleotide probes (20-40 nucleotides long) will be prepared with minimal codon redundancy. These probes will be used to screen the Lambdagt 11-cDNA library and to isolate and sequence the cDNa for the androgen receptor mRNA. The cDNA will then be used to isolate the genomic fragments for the androgen receptor gene(s) and identify the promoter region(s). The cDNA for this hamster androgen receptor mRNA will also be utilized to screen a Lambdagt 11-cDNA library made to human benign prostate hyperplasia messenger RNA. The cDNAs will then be used to study androgen receptor gene expression in growth and differentiation in both normal and abnormal tissues and in cells in a variety of androgen responsive systems.
