Dissecting the function of MYEF2 in neural stem cells and glioblastoma
Item statusRestricted Access
Embargo end date27/06/2021
McCarten, Katrina Anne
Glioblastoma (GBM) is the most common malignant primary brain tumour in adults, with a median survival time of approximately ~15 months after diagnosis. GBM stem cells (GSCs) underpin GBM’s aggressiveness, resistance to radiation and chemotherapy, and disease recurrence. Glioblastoma stem cells (GSCs) are capable of tumour formation upon orthotopic transplantation into immunocompromised mice and share many properties of neural stem cells (NSCs). One feature common to both GSCs and NSCs is high expression of the transcription factor SOX2. SOX2 is known to be essential for GCS self-renewal. To understand further how SOX2 exerts its function in GSCs, the Pollard laboratory studied the interactome of SOX2. Using SICAP-MS – a method that looks at the on-chromatin interactors of the protein of interest – the Pollard laboratory have identified MYEF2 as a candidate interacting partner. MYEF2 contains multiple RNA binding domains and is expressed primarily in the brain and testis. Here I explored the function of MYEF2 within NSCs and GSCs. I confirm Myef2 is expressed within the brain, however was not specifically enriched in stem cell containing regions. CRISPR/Cas9 mediated knock-in of mCherry and HA to Myef2 was successfully carried out in mouse NSCs. We observed that MYEF2 is a nuclear protein which retains nuclear localisation and expression in both proliferative and quiescence NSCs, as well as differentiating progeny. During mitosis, unlike SOX2, MYEF2 is not retained on the mitotic chromatin indicating it is not a “bookmarking” factor. CRISPR/Cas9 mediated knock-out of Myef2 suggests that although it is not essential for the continued proliferation of GSCs, it has a role in regulating the exit from quiescence. Consistent with this, in Myef2 knock-out mouse GSCs, the rate of tumour progression is slower, and mice have a significant survival advantage, suggesting there is an important role for Myef2 in driving tumour growth. In this thesis, I also describe the use of SMASh tag degron technology to precisely control the degradation of SOX2 in mouse NSCs. SMASh, a drug degradable selfcleaving degron, was fused to the C-terminal of endogenous SOX2 in mouse NSCs. We find that this works well and can therefore be a powerful tool in future studies of GSCs.