Gene duplication gives rise to a new 17-kilodalton lipocalin that shows epididymal region-specific expression and testicular factor(s) regulation

Using transgenic mice, we have recently shown that 5 kb of the 5'-flanking region of the mouse epididymal retinoic acid-binding protein (mE-RABP) gene contains all of the information required for spatial and temporal gene exp ression in the epididymis. To identify the important cis-DNA regulatory ele ment(s) involved in the tissue-, region-, and cell-specific expression of t he mE-RABP gene, the 5-kb DNA fragment was sequenced. A computer analysis o f the nucleotide sequence showed the presence of a new gene located 1.7 kb upstream from the mE-RABP gene transcription initiation site. The analysis of the open reading frame showed that the new gene encoded a putative 17-kD a lipocalin (named mEP17) related to mE-RABP. A 600-bp complementary DNA en coding mEP17 was cloned by rapid amplification of 3'-cDNA ends from epididy mal total RNA. Two mEP17 RNA species (1 and 3.1 kb in size) were detected b y Northern blot in the epididymis, but not in other tissues tested. In situ hybridization analyses showed that, unlike mE-RABP messenger RNA (mRNA), w hich is expressed in the distal caput epididymidis, mEP17 mRNA was detected only in the principal cells of the initial segment. The spatial expression and homology with mE-RABP suggest that mEP17 may act as a retinoid carrier protein within the epididymis. mEP17 mRNA expression disappeared 5 days po stcastration. Four days after unilateral castration, mEP17 mRNA had nearly disappeared in the epididymis from the castrated side, but not from the int act side. In addition, testosterone replacement to bilaterally castrated mi ce failed to restore gene expression. We conclude that mEP17 gene expressio n is dependent on testicular factors circulating in the luminal fluid. Toge ther our results suggest that mE-RABP and mEP17 genes were generated by dup lication and that evolution led to a different region-specific gene express ion and regulation in the epididymis.