In vivo analyses of the LHX3 and LHX4 pituitary transcription factors
Abstract
LHX3 and LHX4 are LIM-homeodomain transcription factors with important roles in pituitary and nervous system development. Mutations in the genes encoding these key regulatory proteins are associated with combined pituitary hormone deficiency (CPHD) diseases in human patients and animal models. The studies presented here were undertaken to better understand the roles that LHX3 and LHX4 play in transcriptional regulation of pituitary ontogeny and to gain insight into the molecular origins of CPHD. In collaboration with the University of Leipzig Hospital for Children & Adolescents and the Brooklyn Maimonides Medical Center, a total of eight novel LHX3 and LHX4 mutations were identified and characterized in unrelated patients presenting with CPHD. In many cases, these mutations were found to decrease the transactivation capacity of LHX3 and LHX4 proteins on established genetic targets through a reduction in DNA binding affinity or a disruption in protein partnering. Molecular analyses of the aberrant LHX3 and LHX4 proteins have revealed new aspects of the structural and functional relationships of these factors in endocrine and nervous system development. Ultimately, this work will aid clinicians in establishing more precise molecular diagnoses and treatment regimens for individuals affected by CPHD. In an effort to complement the aforementioned loss-of-function studies, a transgenic approach was undertaken to investigate the individual roles of the LHX3 isoforms in anterior pituitary development and regulation. The LHX3 gene generates at least three structurally distinct protein isoforms, LHX3a, LHX3b, and M2-LHX3. The mRNAs that encode these isoforms maintain unique temporal and spatial expression patterns and, importantly, encode proteins with different abilities to bind and transactivate pituitary target genes in vitro. The alpha glycoprotein subunit (αGSU) promoter was used to drive overexpression of human cDNAs encoding LHX3a, LHX3b, and M2-LHX3 in transgenic mice throughout the developing anterior pituitary. In the adult rodent, expression of the transgene becomes restricted to the gonadotrope and thyrotrope cell lineages. The αGSU-LHX3a and αGSU-LHX3b pedigrees exhibited a sex-specific reduction in viability and displayed reproductive diseases associated with hormonal dysregulation. These gain of function studies provide novel insights to the regulatory properties of distinct LHX3 isoforms and highlight the threshold sensitivity of the anterior pituitary to LHX3 gene dosage.
Degree
Ph.D.
Advisors
Rhodes, Purdue University.
Subject Area
Molecular biology
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