An Otx-related homeodomain protein binds an LHbeta promoter element important for activation during gonadotrope maturation.
The hormone-secreting cell types of the anterior pituitary differentiate in a specific spatial and temporal manner. The alpha-subunit of the glycoprotein hormones appears at embryonic d 11.5 in the mouse, followed by steroidogenic factor-1, which distinguishes the gonadotrope progenitor cells, around embryonic d 14. Gonadotrope maturation is marked by the onset of LHbeta-gene expression 2 d later. The alphaT3-1 and LbetaT2 immortalized mouse pituitary cell lines correspond to these later sequential stages of gonadotrope differentiation. In addition to the early markers of the gonadotrope lineage present in alphaT3-1 cells, LbetaT2 cells also express markers of a mature gonadotrope, including LHbeta and FSHbeta. Using transient transfections to compare expression among gonadotrope and nongonadotrope-derived cell types, we show that the rat 1.8-kb LHbeta promoter directs reporter gene expression specifically to the mature gonadotrope LbetaT2 cell line. Promoter truncation and mutagenesis analyses indicate that the homeodomain (HD) element located at approximately -100 bp relative to the transcriptional start site is essential for this selectivity to LbetaT2 cells when compared with alphaT3-1 cells. In EMSAs, this HD site binds a protein present in LbetaT2 but not other gonadotrope-derived cells. Antibody supershift and competition experiments indicate that this LbetaT2 nuclear protein is a K50 HD protein related to the Otx family, though it is not a known pituitary homeobox transcription factor protein. These studies indicate a role for a novel Otx-related HD protein in gonadotrope maturation during development.
Rosenberg
Suzanne B
SB
Department of Reproductive Medicine, University of California, San Diego, La Jolla, California 92093-0674, USA.
Mellon
Pamela L
PL
D000818:Animals; D001483:Base Sequence; D002454:Cell Differentiation; D002460:Cell Line; D016384:Consensus Sequence / Q000235:genetics; D024202:Electrophoretic Mobility Shift Assay; D006066:Gonads / Q000166:cytology* / Q000378:metabolism*; D006801:Humans; D037101:Luteinizing Hormone, beta Subunit / Q000235:genetics*; D046911:Macromolecular Substances; D051379:Mice; D011401:Promoter Regions, Genetic / Q000235:genetics*; D051381:Rats; D020218:Response Elements / Q000235:genetics; D015533:Transcriptional Activation; D014162:Transfection
D037101:Luteinizing Hormone, beta Subunit; D046911:Macromolecular Substances