Regulation of genome folding by MLL complexes during mouse pluripotent stem cell differentiation
Department: Stem Cell Institute
Supervisor:Dr. Srinjan Basu
Second Supervisors DepartmentWellcome Trust Sanger Institute
The generation of diverse cell types is a hallmark of metazoan biology, yet the mechanisms that initiate and subsequently stabilise cell type-specific gene expression programmes remain ill-defined. Alterations in three-dimensional genome architecture occur during these cell fate transitions but it is still unclear how these rearrangements, which occur alongside changes in transcription, influence differentiation.
These changes are regulated by large protein complexes such as the MLL (mixed lineage leukemia) complexes. How these complexes work is still poorly understood, but they play an important role in fine-tuning gene expression during mammalian development. Mutations in MLL complexes lead to early developmental disorders such as childhood cancer.
We are currently studying how MLL complexes influence genome architecture and transcription in ES cells as they differentiate into neuronal lineages. The aim of this project will be to dissect the role of specific MLL complexes in controlling genome structure and transcription during this process.
To address this, single-cell approaches are key because there is considerable cell heterogeneity in gene expression during this process. Studying the role of these complexes therefore requires examining how they drive genome folding at the level of a single cell. In recent years, we have implemented:
1) a chromosome conformation capture approach called single-cell Hi-C to study genome folding in individual PSCs.
2) 2D and 3D live-cell super-resolution imaging approaches capable of localising single proteins at <15 nm resolution to understand how MLL complexes influence genome folding.
Based on the interest of the student, the project can therefore involve some of the following techniques:
1. ES cell culture (CRISPR/Cas9 cell line generation, Western blots, qPCRs)
2. Hi-C, ChIP-seq, RNA-seq protocols at the bulk or single-cell level
3. 2D/3D super-resolution imaging of one or multiple proteins
4. Computational analysis of ChIP-seq/RNA-seq/Hi-C structures/imaging
Misregulation of MLL complexes often lead to cancer progression making them key drug targets. Imaging assays are also often easily converted into diagnostic tools or drug testing. There is therefore potential for industrial collaborations that can be explored during the project.