Abstract 308 Pulmonary Development Platform, Saturday, 5/1

We are interested in genes that regulate lung development in late gestation. Differential display-PCR was used to identify glucocorticoid-inducible genes in fetal rat lung. A screening of 25% of expressed genes in fetal rat lung cell cultures identified a panel of cDNAs probes representing mRNA whose pattern of developmental and hormonally modulated expression is coordinate with the onset of surfactant synthesis. A day 20 glucocorticoid-treated fibroblast cDNA library was screened using a single probe to isolate the 3.1kb cDNA, LGL1 (AF109674), encoding a deduced polypeptide of 188 amino acids. Northern blot analysis confirmed that LGL1 is induced by glucocorticoid, is differentially expressed in fetal lung adjacent fibroblasts (detected at d16, maximally at d21) but not detected in lung epithelium. LGL1 is also detectable, albeit at lower levels, in rat fetal heart, kidney, intestine, as well as in adult human and mouse lung. Northern analysis also provided preliminary evidence that LGL1 expression is decreased at fetal day 18 in homozygous GR knockout mice (mice lacking the glucocorticoid receptor) when compared to wild-type or heterozygous mice. RT-PCR analysis of human fetal tissues detected LGL1 expression in lung at 12, 16 and 18 weeks gestation, in gut at 12 and 18 weeks, and in kidney, heart and CNS at 18 weeks. In situ hybridization studies demonstrated that LGL1 expression (fetal rat lung, kidney and gut) is restricted to the mesenchyme. LGL1 demonstrates 68% identity (81% homology) to P25TI, a polypeptide with weak trypsin inhibitor activity, identified in human glioblastoma and neuroblastoma cells but not detected in normal human tissues. Both LGL1 and P25TI belong to the CRISP family of androgen regulated, cystine rich secretory proteins. Trypsin is produced by both normal bronchial epithelial cells and in adenocarcinoma of the lung. Like other proteases, it is believed to be important in the tight control of extracellular matrix degradation that is critical to cell and organ development. We have expressed a his-tagged LGL1 in the pRSET E. coli system to further characterize its role in fetal lung maturation. In common with other regulatory proteins that play a role in development and oncogenic processes, we speculate that LGL1 may have a role in normal development via the regulation of extracellular matrix degradation.

Funded by an Medical Research Council of Canada Grant to F. Kaplan and a McGill University/Montreal Children's Hospital Research Institute Studentship to F. Q. Kassamali.