Regulation of gene-expression by the transcription factor Pax6
Our research is focused on the transcription factor Pax6, with the overall aim of detecting new target genes in order to understand Pax6’s role in normal development and cancer.
Pax6 is an evolutionary conserved transcription factor, expressed in the central nervous system, eye, nose and pancreas during vertebrate embryo development. Pax6 heterozygous mutants lead to the eye phenotype Aniridia in humans, and are also coupled to glucose intolerance and the lack of pineal gland. Several cancer cell lines show deregulated expression of Pax6. None of the published Pax6 targets genes are able to fully explain the phenotypes observed when Pax6 is mutated, or what Pax6 possible could contribute to in tumors.
To identify new target genes we use:
- Chromatin immunoprecipitation (ChIP) and microarray of; 1) genetically engineered Pax6 inducible cell lines and 2) cancer cell lines endogenously expressing Pax6 treated with Pax6 siRNA. A combination of these two methods (ChIP-on-chip) have also been tried out.
- Co-injection of ChIP selected elements and enhancer reporter assays in zebrafish , as well as in-situ hybridization of a putative Pax6 target gene in Pax6.1 and Pax6.2 morpholino treated zebrafish embryos.
- 2D gel-electrophoresis, Mass-spechtrometry and SILAC on protein extracts from stably transfected cell-lines that show morphological changes upon Pax6 and Pax6(5a) expression.
Verification of the most interesting target genes are done by electromobility shift assay (EMSA), as well as RT-PCR, qPCR and transient transfection/reporter-gene assay.
Recently we have also started a collaboration project on “Safety Pharmacology” with the School of Pharmacy in Oslo. The main aim of this project is to test the feasibility of chicken and zebrafish as models in CNS safety pharmacology tests for drugs relevant in pediatric medicine or pregnant mothers. Glucocorticoids, anti-depressants and anti-epileptics will be tested.
Group members 2011:
Ingvild Mikkola (Group leader, Associate professor)
Rune Hogseth (Technician)
Yury Kiselev (MD, PhD student)
Siri Forsdahl (Cand. Pharm, PhD student)
Beate Hegge (Technician)
Mina Mobasheri (Master student parmacy)