The anterior pituitary contains six phenotypically distinct cell types, each of which produces a different peptide hormone in a cell-specific manner. Pituitary lactotroph cells specialize in prolactin (PRL) gene expression, which is mediated by multiple, homologous promoter elements that bind a single lactotroph-specific factor, LSF-1. We have identified a highly conserved DNA hexamer motif within the lactotroph-specific DNA elements (LSE), and purified LSF-1 via LSE DNA-affinity chromatography. LSF-LSF-1 is distinct from GHF-1, which mediates GH gene expression in somatotrophs. Also, we've mapped a negative regulatory element that inhibits rPRL transcription in nonpituitary cells, and identified F2F as the factor which binds to this site. Cell-type restriction may be due to tissue-specific nu-clear proteins (trans-acting factors) which activate selective genetic programs by interacting with short DNA promoter elements (cis-active sequences). Despite this significant progress in our understanding of gene regulation, the precise mechanisms by which factor occupancy of cis-active elements actually governs gene transcription remains unknown.
The specific aims of this proposal are to identify the exact DNA sequence and distance requirements of the LSEs, and to reconstitute lactotroph-specific transcription using affinity-purified, recombinant, or synthetic) will be added to nonpituitary (HeLa) cell-free extracts to reconstitute pituitary-specific rPRL gene expression. Additionally, the effects of pre-treating LSF-1 with phosphatase, and kinases A & C, will be evaluated. Short synthetic LSF-1 oligopeptides of defined function previously determined by cell-free transcription will be used for preparation of domain-specific antibodies. Synthetic LSF-1 oligopeptides will be used to prepare an LSF-1 protein-affinity column to purify other pituitary nuclear factors that specifically interact with LSF- 1. These will be analyzed by SDS PAGE, and tested for function in the cell-free transcription assay. Finally, the role of LSF-1 phosphorylation in vivo will be evaluated by combining antibody precipitation and metabolic labelling studies. The development of molecular probes, with in vivo gene transfer techniques provide extremely sensitive methods to accomplish these aims. Finally, these studies set the stage for future work aimed at defining the role of LSF-1 & F2F in normal and abnormal pituitary development.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Research Project (R01)
Project #
Application #
Study Section
Endocrinology Study Section (END)
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
University of Colorado Denver
Schools of Medicine
United States
Zip Code
Tong, Yunguang; Zhou, Jin; Mizutani, Jun et al. (2011) CEBPD suppresses prolactin expression and prolactinoma cell proliferation. Mol Endocrinol 25:1880-91
Jonsen, Matthew D; Duval, Dawn L; Gutierrez-Hartmann, Arthur (2009) The 26-amino acid ss-motif of the Pit-1ss transcription factor is a dominant and independent repressor domain. Mol Endocrinol 23:1371-84
Jedlicka, Paul; Sui, Xiaomei; Gutierrez-Hartmann, Arthur (2009) The Ets dominant repressor En/Erm enhances intestinal epithelial tumorigenesis in ApcMin mice. BMC Cancer 9:197
Jedlicka, Paul; Sui, Xiaomei; Sussel, Lori et al. (2009) Ets transcription factors control epithelial maturation and transit and crypt-villus morphogenesis in the mammalian intestine. Am J Pathol 174:1280-90
Jedlicka, Paul; Gutierrez-Hartmann, Arthur (2008) Ets transcription factors in intestinal morphogenesis, homeostasis and disease. Histol Histopathol 23:1417-24
Ferry, A L; Locasto, D M; Meszaros, L B et al. (2005) Pit-1beta reduces transcription and CREB-binding protein recruitment in a DNA context-dependent manner. J Endocrinol 185:173-85
Farrow, Kathryn N; Bradford, Andrew P; Tentler, John J et al. (2004) Structural and functional analysis of the differential effects of c-Jun and v-Jun on prolactin gene expression. Mol Endocrinol 18:2479-90
Dunkelberg, J C; Gutierrez-Hartmann, A (2001) LZ-FYVE: a novel developmental stage-specific leucine zipper, FYVE-finger protein. DNA Cell Biol 20:403-12
Diamond, S E; Gutierrez-Hartmann, A (2000) The Pit-1beta domain dictates active repression and alteration of histone acetylation of the proximal prolactin promoter. J Biol Chem 275:30977-86
Farrow, K N; Gutierrez-Hartmann, A (1999) Transforming growth factor-beta1 inhibits rat prolactin promoter activity in GH4 neuroendocrine cells. DNA Cell Biol 18:863-73

Showing the most recent 10 out of 35 publications