Rabbit lung surfactant protein A gene: identification of a lung-specific DNase I hypersensitive site

Expression of the gene encoding pulmonary surfactant protein A, SP-A, is lung specific and developmentally and hormonally regulated. Previously, we observed that SP-A gene transcription is initiated in fetal rabbit lung after day 21 of gestation and reaches maximal levels by day 28. In the present study, a cDNA specific for rabbit SP-A was used to isolate the SP-A gene from a rabbit genomic library. A 7.6-kb fragment containing the entire structural gene and approximately 380 bp of 5'-flanking DNA was isolated and characterized. The transcription initiation site, mapped by primer extension analysis, was localized 23 bp downstream of a putative TATA element. The structural gene is composed of five exons and four introns. The first exon encodes the 5'-untranslated region of the mRNA; the translation initiation site is in exon II, and exon V contains the two polyadenylation sites that give rise to the 2.0- and 3.0-kb species of SP-A mRNA. A potential adenosine 3',5'-cyclic monophosphate (cAMP)-regulatory element (CRE) was identified at -261 bp, and sequences with homology to glucocorticoid-regulatory element (GRE) half-sites were found at -150 and -190 bp upstream of the transcription initiation site and within the first intron. A DNase I hypersensitive site was identified in genomic DNA isolated from 21- and 28-day fetal and adult rabbit lung tissues. This site was mapped within the 5'-flanking region of the SP-A gene, at approximately -80 to -180 bp upstream of the transcription initiation site. The absence of this hypersensitive site in genomic DNA of liver, kidney, and heart tissues suggests that altered chromatin structure may serve a role in lung-specific SP-A gene expression. The presence of this tissue-specific DNase I hypersensitive site in lung nuclei from 21-day gestational age fetal rabbits suggests that the SP-A gene may exist in an accessible conformation prior to the time of transcription initiation.