DNA repair by HDR in experimental tumorigenesis
The aim of this thesis was to further understand how defects in the homology-directed repair (HDR) pathway affect tumor formation and development. To study proteins involved in HDR is challenging since most of its members are required for cell viability. For our studies we have therefore taken a dominant-negative approach to address the importance of the BRCA1 interacting protein BARD1 in HDR (paper I) and investigate the role of HDR in PDGFB-induced gliomagenesis by interfering with a key protein, RAD51 (paper II). In addition we have also investigated how Platelet-derived growth factor A receptor (PDGFRA) is transcriptionally regulated by interleukin-1b (IL-1b) (paper III).
The breast cancer susceptibility gene BRCA1 is frequently mutated in hereditary breast and ovarian cancers. BRCA1 has been implicated in many different cellular processes, among them DNA repair by HDR. In Paper I we investigated if BARD1 was involved in HDR through its interaction with BRCA1. We could show that expression of a truncated BARD1 decreased HDR of an induced double-strand break and that this decrease was even more pronounced in Brca1-deficient cells expressing a splice variant of Brca1 that still are able to bind BARD1. We could also show that the role of BARD1 in HDR was dependent on binding to BRCA1 and that the HDR defect resulting from the truncated BARD1 was caused by lack of regulatory elements in the C-terminal end of BARD1 and not caused by improper cellular localization of either the BARD1 construct or the endogenous Brca1 protein. We conclude that BARD1 is important for HDR and that the repair function of BARD1 is dependent on its interaction with BRCA1.
The expression of PDGFRa is strictly regulated during embryogenesis and aberrant PDGFRa expression can lead to malignant transformation, in brain tumors for example. Highly malignant gliomas are the most frequent primary tumor of the central nervous system in adults. To study the effect of different genetic changes in gliomagenesis, glioma-like tumors can be induced by intracerebral injections of oncogene carrying retroviruses. The RCAS/tv-a model system provides the possibility to study combinations of different genetic alterations in a specific cell type. In paper II we have used wild type and Arf-/- nestin tv-a (Ntv-a) transgenic mice, where expression of the RCAS constructs were directed to neural progenitor cells. It has previously been shown that expression of PDGFB can induce glioma-like tumors in these mice.
To investigate the role of HDR in tumorigenesis we co-expressed RAD51 or a DNA repair deficient RAD51 (RAD51KR) contruct with PDGFB in either wild type or Arf-/- Ntv-a mice. We could show that co-expression of RAD51 or RAD51KR can suppress PDGFB-induced tumorigenesis in wild type mice. However, only RAD51 was able to suppress tumor formation in the Arf-null background. We could also show that all the PDGFB-induced tumors were aneuploid, independent of genotype, tumor grade or tumor size. Interestingly, expression of RAD51 or RAD51KR reduced aneuploidy in the PDGFB-induced tumors.
List of scientific papers
I. Westermark UK, Reyngold M, Olshen AB, Baer R, Jasin M, Moynahan ME (2003). BARD1 participates with BRCA1 in homology-directed repair of chromosome breaks. Mol Cell Biol. 23(21): 7926-36.
https://pubmed.ncbi.nlm.nih.gov/14560035
II. Westermark UK, Forsberg N, Bråsäter D, Helgadottir HR, Eriksson A, Zetterberg A, Jasin M, Nistér M, Uhrbom L (2008). RAD51 suppresses PDGFB-induced gliomagenesis and genetic instability in wild type and Arf-/- mice. [Submitted]
III. Afink G, Westermark UK, Lammerts E, Nistér M (2004). C/EBP is an essential component of PDGFRA transcription in MG-63 cells. Biochem Biophys Res Commun. 315(2): 313-8.
https://pubmed.ncbi.nlm.nih.gov/14766209
History
Defence date
2008-06-05Department
- Department of Oncology-Pathology
Publication year
2008Thesis type
- Doctoral thesis
ISBN
978-91-7409-002-4Number of supporting papers
3Language
- eng