The goal of our research is to define the molecular mechanisms involved in lung type II cell-specific, developmental and hormonal regulation of surfactant synthesis in fetal lung. To achieve this goal, we have focused on the gene encoding surfactant protein-A (SP-A), a major surfactant protein expressed primarily in type II pneumonocytes and is developmentally regulated in concert with surfactant glycerophospholipid synthesis. SP-A gene transcription in midgestation rabbit and human fetal lung in culture is induced by hormones and factors that increase cyclic AMP (cAMP). In deletion mapping and mutagenesis studies, we have found that basal and cAMP induction of rabbit and human SP-A promoter activity is critically dependent upon cooperative interactions of at least four identified DNA response elements which lie within 400 bp upstream of the SP-A transcription initiation site. Mutagenesis of any one of these elements has a marked effect to reduce basal and cAMP induction of SP-A promoter activity. In studies using transgenic mice carrying SP-A:human growth hormone (hGH) reporter genes, we have found that as little as 400 bp of SP-A 5'-flanking sequence mediates lung- specific and appropriate developmental regulation of hGH expression. In the proposed research, we will use transgenic mice for deletion mapping and mutagenesis to further define critical response elements, and for a promoter building strategy, whereby tissue/cell-specific and developmental regulation of transgene expression will be analyzed in mice carrying SP-A:hGH fusion genes containing the SP-A basal promoter fused downstream of the identified response elements, either individually or in various combinations. The functional role of these elements within the endogenous mouse SP-A gene will be analyzed by gene targeting. Transcription factors (TFs) that bind to these and other identified response elements will be isolated by expression screening or yeast one-hybrid system. Upon identification of TFs crucial for SP-A gene regulation, we will explore effects of cAMP on their DNA binding and transcriptional activities, their interactions with each other and with co-activators, and their functional role in mice by gene targeting or dominant-negative inactivation. Finally, in light of the important role of cAMP in type II cell differentiation and the regulation of SP-A gene expression, we will analyze developmental changes in expression and subcellular localization of protein kinase A (PKA) regulatory (R) and catalytic (c) subunits and of anchor proteins important for PKA nuclear localization, during lung development and type II cell differentiation. To assess the role of PKA in lung development and surfactant synthesis, we will create transgenic mice carrying a dominant negative R subunit under control of SP-A and SP-C promoters to inhibit PKA activity in a type II cell-specific, developmentally timed manner.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37HL050022-08
Application #
6183524
Study Section
Human Embryology and Development Subcommittee 1 (HED)
Project Start
1993-04-01
Project End
2003-03-31
Budget Start
2000-04-01
Budget End
2001-03-31
Support Year
8
Fiscal Year
2000
Total Cost
$341,423
Indirect Cost
Name
University of Texas Sw Medical Center Dallas
Department
Biochemistry
Type
Schools of Medicine
DUNS #
City
Dallas
State
TX
Country
United States
Zip Code
75390
Maitra, Meenakshi; Wang, Yongyu; Gerard, Robert D et al. (2010) Surfactant protein A2 mutations associated with pulmonary fibrosis lead to protein instability and endoplasmic reticulum stress. J Biol Chem 285:22103-13
Mendelson, Carole R (2009) Minireview: fetal-maternal hormonal signaling in pregnancy and labor. Mol Endocrinol 23:947-54
Liu, Dongyuan; Benlhabib, Houda; Mendelson, Carole R (2009) cAMP enhances estrogen-related receptor alpha (ERRalpha) transcriptional activity at the SP-A promoter by increasing its interaction with protein kinase A and steroid receptor coactivator 2 (SRC-2). Mol Endocrinol 23:772-83
Liu, Dongyuan; Yi, Ming; Smith, Margaret et al. (2008) TTF-1 response element is critical for temporal and spatial regulation and necessary for hormonal regulation of human surfactant protein-A2 promoter activity. Am J Physiol Lung Cell Mol Physiol 295:L264-71
Islam, Kazi Nazrul; Mendelson, Carole R (2008) Glucocorticoid/glucocorticoid receptor inhibition of surfactant protein-A (SP-A) gene expression in lung type II cells is mediated by repressive changes in histone modification at the SP-A promoter. Mol Endocrinol 22:585-96
Liu, Dongyuan; Hinshelwood, Margaret M; Giguere, Vincent et al. (2006) Estrogen related receptor-alpha enhances surfactant protein-A gene expression in fetal lung type II cells. Endocrinology 147:5187-95
Islam, Kazi Nazrul; Mendelson, Carole R (2006) Permissive effects of oxygen on cyclic AMP and interleukin-1 stimulation of surfactant protein A gene expression are mediated by epigenetic mechanisms. Mol Cell Biol 26:2901-12
Alcorn, Joseph L; Islam, Kazi N; Young, Pampee P et al. (2004) Glucocorticoid inhibition of SP-A gene expression in lung type II cells is mediated via the TTF-1-binding element. Am J Physiol Lung Cell Mol Physiol 286:L767-76
Gao, Erwei; Wang, Ying; Alcorn, Joseph L et al. (2003) Transcription factor USF2 is developmentally regulated in fetal lung and acts together with USF1 to induce SP-A gene expression. Am J Physiol Lung Cell Mol Physiol 284:L1027-36
Islam, Kazi Nazrul; Mendelson, Carole R (2002) Potential role of nuclear factor kappaB and reactive oxygen species in cAMP and cytokine regulation of surfactant protein-A gene expression in lung type II cells. Mol Endocrinol 16:1428-40

Showing the most recent 10 out of 18 publications