Analysis of tissue- and hormone-specific regulation of the human prolactin-inducible protein/gross cystic disease fluid protein-15 gene in transgenic mice.

The human prolactin-inducible protein/gross cystic disease fluid protein-15 (PIP/GCDFP-15) gene is expressed in more than 90% of human breast cancer biopsies but not in the normal mammary gland. However, it is expressed in several normal human apocrine glands such as the lacrimal and salivary glands. In human breast cancer cell lines, the gene is regulated by a number of hormones including androgen and prolactin. It is not known whether gene expression in normal tissues is under similar hormonal control. To understand the mechanisms by which hormone- and tissue-specific expression of the human PIP/GCDFP-15 gene are regulated in vivo, we generated transgenic mice using a 13.7 kb genomic DNA fragment containing the entire 7 kb human gene, together with 2.9 kilobases of 5' and 3.8 kilobases of 3' flanking sequences. The human PIP/GCDFP-15 transgene was found to be expressed in both the lacrimal and salivary glands but was not expressed in the mammary glands of transgenic mice. This tissue-specific pattern of the transgene expression in the mouse was very similar to that of the endogenous human PIP/GCDFP-15 gene, and to the endogenous mouse,gene. In the mouse salivary glands, the transgene expression was highest in the parotid, considerably less in the submaxillary (submandibular) and absent in the sublingual glands. In the mouse lacrimal gland, as in the human breast cancer cell lines, the human PIP/GCDFP-15 transgene was also up-regulated by androgen. These studies demonstrate that the human gene with its 6.3 kb flanking sequences is able to confer gene expression in vivo in a tissue-specific and hormone-responsive manner.

A new member of the hormonally regulated rodent submaxillary gland glycoprotein gene family: cDNA cloning and tissue specific expression.

Polymerase chain reaction was used to amplify and identify two related rat submaxillary gland glycoprotein (rSMGGP and rSMGGP1) cDNAs. They were 489 bp and 594 bp long respectively. The shorter cDNA (rSMGGP) was identical to the previously published rat spot-I protein. The longer cDNA (rSMGGP1) had an additional (117 bp) unique nucleotide sequence in the 3' coding region, and the overall homology between the two cDNAs was 78%. rSMGGP also had a 68% homology to the mouse submaxillary gland glycoprotein (mSMGGP) cDNA. The predicted translated product of rSMGGP1 was 130 amino acids long, 39 amino acids longer than the rSMGGP. The region of greatest diversity between the putative peptides of the two rat cDNAs and the mouse cDNA was in the carboxy terminus. Northern blot analysis, using both rat cDNAs as probes, showed hybridization to an mRNA transcript (650 bases) in the submaxillary and lacrimal gland of the normal adult male and female rat. A larger transcript (approximately 700 bases) was induced under conditions of altered hormonal profiles: hypophysectomy, pregnancy/lactation, and castration. Dihydrotestosterone administration inhibited expression of the two transcripts in both the lacrimal and submaxillary glands of male and female rats. The labelled 117 bp DNA fragment unique to the rSMGGP1 cDNA hybridized only to the 700 base transcript in the rat lacrimal and submaxillary gland suggesting that differential exon usage produces the two variant mRNAs. The regulation of the SMGGP gene expression may provide yet another useful model for studying the mechanism of down-regulation of genes by androgen and the identification of tissue specific factors in the lacrimal and submaxillary gland.

Tissue-specific androgen-inhibited gene expression of a submaxillary gland protein, a rodent homolog of the human prolactin-inducible protein/GCDFP-15 gene.

The human PRL-inducible protein (PIP)/gross cystic disease fluid protein-15 is expressed in pathological conditions of the mammary gland and in several exocrine tissues, such as the lacrimal, salivary, and sweat glands. In human breast cancer cells, the expression of PIP/gross cystic disease fluid protein-15 is stimulated by androgen and PRL, and inhibited by estrogen. However, it is not known whether the expression of PIP in other tissues is under similar hormonal regulation. In the present study we employed reverse transcriptase-polymerase chain reaction followed by rapid amplification of complementary DNA (cDNA) ends to amplify the PIP cDNA homolog, the submaxillary gland protein (SMGP) in the mouse. The mouse PIP/SMGP cDNA encodes a putative secreted peptide of 144 amino acids with a 51% identity with human PIP. Using the mouse PIP/SMGP cDNA as a probe, we examined the tissue- and cell-specific expression of PIP/SMGP messenger RNA by in situ hybridization and Northern blot analysis of mouse and rat tissues. Hormonal regulation was also studied in the rat. PIP/SMGP messenger RNA expression was only detected in the lacrimal and submaxillary glands of the rodents. In the rat submaxillary gland, PIP/SMGP gene expression was confined to the acinar cells. In the male rat lacrimal gland, castration resulted in an increase in expression, and in both male and female rats, androgen replacement abolished PIP/SMGP gene expression. This pattern of regulation was not observed in the submaxillary gland and was actually reversed in human breast cancer cells. PRL had no effect on the regulation of PIP/SMGP in either salivary or lacrimal glands. Our study indicates that tissue-specific factors are important in determining the hormone responsiveness of the PIP/SMGP gene. Regulation of the PIP/SMGP gene in vivo may provide a useful model system to study the mechanism of down-regulation of expression by androgen in a tissue-specific manner.