Filamin A is a well-characterised cytoskeletal protein which regulates cell shape and migration by cross-linking with actin. Filamin A mutations cause a number of human developmental disorders, many of which exhibit skeletal dysplasia. However, the molecular mechanisms by which Filamin A affects skeletal development are unknown. The transcription factor Runx2 is a master regulator of osteoblast and chondrocyte differentiation. Data presented in this thesis show that Filamin A forms a complex with Runx2 in osteoblastic cell lines. Moreover, it is demonstrated that Filamin A is present in the nucleus in several cell lines, including those of osteoblastic origin. The data presented show that the Filamin A/Runx2 complex suppresses the expression of the gene encoding the matrix-degrading enzyme, matrix metalloproteinase-13 (MMP-13), which is an important osteoblastic differentiation marker. ChIP assays were employed to demonstrate that endogenously expressed Filamin A associates with the promoter of the MMP-13 gene. In addition, Filamin A is not only located in the nucleus but also in the nucleolus, an important nuclear compartment involved in ribosomal RNA (rRNA) transcription. Ribosomal DNA promoter-driven reporter assays, Filamin A-knockdown experiments and exogenous Filamin A transfections demonstrated that Filamin A and Runx2 can repress ribosomal gene expression activity. Importantly, Filamin A is recruited to the human ribosomal DNA promoter, suggesting its direct involvement in the regulation of rRNA transcription. These findings reveal a novel role of Filamin A in the direct regulation of ribosomal gene expression. Finally, by using microarray technology, changes in gene expression were identified when Filamin A was downregulated. Some of the differentially expressed genes were known orchestrators of bone development. The data presented in this thesis strengthen the link between Filamin A and bone development and provide a molecular rationale for how Filamin A, acting as a regulator of gene expression, might influence osteoblastic differentiation.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:538413 |
| Date | January 2011 |
| Creators | Lopez Camacho, Cesar |
| Contributors | Shore, Paul |
| Publisher | University of Manchester |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | https://www.research.manchester.ac.uk/portal/en/theses/a-new-role-for-filamin-a-as-a-regulator-of-runx2-function(88321064-5c82-4f2b-a755-911ed3b42b2e).html |
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