This is a renewal application requesting five years of funding to continue Dr. Camper s research program to positionally clone and functionally characterize the mouse Ames dwarf (df) gene. The df mutation arose in descendants of an X-irradiated male mouse at Iowa State University in Ames, Iowa. Df is a recessive mutation that produces severe proportional dwarfism, hypothyroidism and infertility. The growth insufficiency results from a lack of thyroid stimulating hormone (TSH) and growth hormone (GH), and a lack of prolactin (PRL) contributes to the infertility of females. The Ames dwarf mice, like the Snell (Pitdw) dwarf mice, exhibit severe hypocellularity due to a deficiency in the three pituitary cells that produce PRL, GH, and TSH: lactotropes, somatotropes, and thyrotropes, respectively. The simultaneous loss of these three cell types suggest that they derive from a common precursor cell, and the similarity of the phenotypes of the non-allelic df and dw mutants suggests that the two mutated genes are involved in the proliferation and/or differentiation of this precursor cell. The dw mutation has been shown to result from defects in the Pit-1 transcription factor gene.
In Specific Aim 1 Camper proposes to clone the df gene. The df critical region will be physically mapped and the 3.5 Mb YAC and P1 contig encompassing the 635 kb df critical region will be completed. Exon trapping and cDNA selection will be applied to identify transcripts within the df critical region. Full length cDNA clones will be generated for the identified transcripts and these clones will be used in Northern analyses, and if necessary RNase protection and RT-PCR assays, to determine the tissue distribution of the cloned transcripts. These studies will allow Camper to prioritize transcripts as products of possible candidate df genes. Candidate gene sequences will then be compared in normal and df mutant mice to detect mutations in the putative df gene.
In Specific Aim 2, Camper will characterize the intron/exon structure and 5 end of the candidate df gene. In addition, df gene expression gene will be characterized by immunostaining of tissue sections, RT-PCR, RNase protection, in situ hybridization, and Northern analysis of pituitary cell lines. Confirmation that a mutation in a candidate gene is responsible for the df phenotype will be obtained in Specific Aim 3 by transgene correction, the generation of gain of function alleles, and the production of loss of function mutations. These studies will also begin to probe df gene function. Camper proposes in Specific Aim 4 to clone and chromosomally map the human df gene and to assess the association of df with human pituitary tumors and families of short stature.
Specific Aim 5 will establish the role of the df gene within the hierarchy of genes regulating pituitary development by comparing the expression of early pituitary markers in df and dw and by examining df gene expression in other mouse mutants with pituitary defects.

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
Research Project (R01)
Project #
5R01HD030428-06
Application #
2673717
Study Section
Mammalian Genetics Study Section (MGN)
Project Start
1993-04-01
Project End
2002-03-31
Budget Start
1998-04-01
Budget End
1999-03-31
Support Year
6
Fiscal Year
1998
Total Cost
Indirect Cost
Name
University of Michigan Ann Arbor
Department
Genetics
Type
Schools of Medicine
DUNS #
791277940
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
Pérez Millán, María I; Vishnopolska, Sebastian A; Daly, Alexandre Z et al. (2018) Next generation sequencing panel based on single molecule molecular inversion probes for detecting genetic variants in children with hypopituitarism. Mol Genet Genomic Med :
Cheung, Leonard Y M; George, Akima S; McGee, Stacey R et al. (2018) Single-Cell RNA Sequencing Reveals Novel Markers of Male Pituitary Stem Cells and Hormone-Producing Cell Types. Endocrinology 159:3910-3924
Cheung, Leonard Y M; Okano, Hideyuki; Camper, Sally A (2017) Sox21 deletion in mice causes postnatal growth deficiency without physiological disruption of hypothalamic-pituitary endocrine axes. Mol Cell Endocrinol 439:213-223
Correa, Fernanda A; França, Marcela M; Fang, Qing et al. (2017) Growth hormone deficiency with advanced bone age: phenotypic interaction between GHRH receptor and CYP21A2 mutations diagnosed by sanger and whole exome sequencing. Arch Endocrinol Metab 61:633-636
Cheung, Leonard Y M; Davis, Shannon W; Brinkmeier, Michelle L et al. (2017) Regulation of pituitary stem cells by epithelial to mesenchymal transition events and signaling pathways. Mol Cell Endocrinol 445:14-26
Fang, Qing; Benedetti, Anna Flavia Figueredo; Ma, Qianyi et al. (2016) HESX1 mutations in patients with congenital hypopituitarism: variable phenotypes with the same genotype. Clin Endocrinol (Oxf) 85:408-14
Davis, Shannon W; Keisler, Jessica L; Pérez-Millán, María I et al. (2016) All Hormone-Producing Cell Types of the Pituitary Intermediate and Anterior Lobes Derive From Prop1-Expressing Progenitors. Endocrinology 157:1385-96
Fang, Qing; George, Akima S; Brinkmeier, Michelle L et al. (2016) Genetics of Combined Pituitary Hormone Deficiency: Roadmap into the Genome Era. Endocr Rev 37:636-675
Mortensen, Amanda H; Camper, Sally A (2016) Cocaine-and Amphetamine Regulated Transcript (CART) Peptide Is Expressed in Precursor Cells and Somatotropes of the Mouse Pituitary Gland. PLoS One 11:e0160068
Pérez Millán, María Inés; Brinkmeier, Michelle L; Mortensen, Amanda H et al. (2016) PROP1 triggers epithelial-mesenchymal transition-like process in pituitary stem cells. Elife 5:

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