Nuclear Factor I/Thyroid Transcription Factor 1 Interactions Modulate Surfactant Protein C Transcription

Abstract

Surfactant protein C (SP-C; Sftpc) gene expression is restricted to pulmonary type II epithelial cells. The proximal SP-C promoter region contains critical binding sites for nuclear factor I (NFI) and thyroid transcription factor 1 (TTF-1; also called Nkx2.1). To test the hypothesis that NFI isoforms interact with TTF-1 to differentially regulate SP-C transcription, we performed transient transfection assays in JEG-3 cells, a choriocarcinoma cell line with negligible endogenous NFI or TTF-1 activity. Cotransfection of NFI family members with TTF-1 induced synergistic activation of the SP-C promoter that was further enhanced by p300. TTF-1 directly interacts with the conserved DNA binding and dimerization domain of all NFI family members in coimmunoprecipitation and mammalian two-hybrid experiments. To determine whether SP-C expression is regulated by NFI in vivo, a chimeric fusion protein containing the DNA binding and dimerization domain of NFI-A and the Drosophila engrailed transcriptional repression domain (NFIen) was conditionally expressed in mice under control of a doxycycline-inducible transgene. Induction of NFIen in a subset of type II cells inhibited SP-C gene expression without affecting expression of TTF-1 in doxycycline-treated double-transgenic mice. Taken together, these findings support the hypothesis that NFI family members interact with TTF-1 to regulate type II cell function.

ACKNOWLEDGMENTS

We thank J. A. Whitsett for providing the Drosophila engrailed transcriptional repression domain, monoclonal TTF-1 antibody, and the CCSP-rtTA activator mice and for helpful discussions and suggestions. We also thank H. Bujard for the rtTA expression plasmid, C. Yan for providing some of the plasmids used in the mammalian two-hybrid analysis, and T. Inoue for the pBETNFIb1f untagged mouse NFI-A expression plasmid. Susan Wert and the Molecular Morphology Core helped with ISH and Kathryn Foss provided expert technical assistance.

This work was supported by NIH grants HL60907 (C.J.B.), HD34908, and DK58401 (R.M.G.) and the Charlotte R. Schmidlapp Women Scholars Award (C.J.B.).