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Title: Regulation of Fanconi Anaemia complementation group a (Fanca) expression by GnRH : a role in reproduction
Author: Larder, R.
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2005
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Differential display RT-PCR (DD-RT-PCR) analysis of GnRH stimulated transcripts has identified that a DNA damage repair gene, Fanconi Anaemia complementation group a (Fanca), is differentially expressed in response to hormone. FANCA is mutated in > 60% of cases of Fanconi anaemia (FA), a genetically heterogeneous autosomal recessive disorder characterised by bone marrow failure, endocrine tissue cancer susceptibility and infertility. Detailed analysis of Fanca mRNA expression using northern blotting, semi-quantitative RT-PCR and quantitative RT-PCR analysis reveals that GnRH induces a rapid and transient increase in Fanca mRNA within LβT2 gonadotroph cells. Indirect immunofluorescence and western blotting analysis show that Fanca protein is expressed in the cytoplasm and nucleus of LβT2 cells and GnRH induces a transient 2-fold increase in Fanca protein levels within both cellular compartments. Furthermore, treatment with inhibitors of nuclear import and export demonstrated that Fanca protein actively shuttles between the nucleus and cytoplasm of gonadotroph cells, via a CRM-1 (yeast chromosome region maintenance protein 1) dependent mechanism. Transient transfection assays using wildtype and dominant negative, point mutated forms of FANCA reveal that wildtype FANCA protein is required for GnRH induced activity of the αGSU promoter, but not LHβ or FSHβ promoter activity. The construction of adenoviral vectors expressing wildtype and mutant FANCA proteins and the characterisation of siRNAs to knockdown expression of Fanca in LβT2 cells will allow the confirmation of a novel role for Fanca in gonadotropin gene transcription. The discovery that GnRH regulates Fanca expression, which in turn regulates GnRH induced αGsu transcription, provides the first molecular evidence of a role for Fanca in the control of fertility.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available