PhD Project - Defining the Molecular and Developmental Basis of MBD5-associated neurodevelopmental disorder (MAND)

Vacancy Reference Number
2022-SIDB-ILLINGWORTH
Closing Date
16 Jan 2022
Address
University of Edinburgh

Background:

Loss of function mutations in chromatin modifier proteins are a prominent cause of neurodevelopmental disorders (NDDs). MBD5-associated neurodevelopmental disorder (MAND) is rare human NDD associated with intellectual disability (ID), motor deficits, seizures and autism1. MAND is caused by mutations in the MBD5 gene, the protein product of which associates with ASXL1,2 or 3 and BAP1 to form the polycomb repressive deubiquitinase (PR-DUB) complex. In mouse ES cells, PR-DUB has been shown to regulate gene expression by controlling global H2A ubiquitination levels2. Despite this mechanistic insight, and an expanding repertoire of causal MBD5 mutations; how mutations in MBD5 give rise to MAND phenotypes is currently unknown.

Rationale & hypothesis:

The formation of the human brain requires a balance between the expansion and differentiation of neural stem cells. This is coordinated, in part, by chromatin modifiers that control developmental gene expression programmes. We hypothesise that the gross developmental deficits associated with MBD5 mutations arise due to defective control of neural stem cell populations. To test this hypothesis, the student will establish and characterise cerebral organoids bearing MAND causal mutations.

Aims:

  • Engineer MAND mutations into isogeneic, early-stage primary human cortical neural stem cells (NSCs; a biologically relevant and genetically tractable cell system derived in the lab of Prof. Steve Pollard - Centre for Regenerative Medicine).
  • Determine the epigenetic and transcriptional landscape of MAND mutant NSCs.
  • Derive cerebral organoids from human induced pluripotent cells (iPSCs) and primary MAND mutant NSCs.
  • Determine the gross and relative (between mutations) developmental impact of MAND mutations using cerebral organoids as a model system.

Use these complementary approaches in genomics3 and neurodevelopmental biology to advance our understanding of the molecular basis of MBD5-associated neurodevelopmental disorder.

Training outcomes:

  • Culture of primary NSCs and in vitro differentiated cerebral organoids.
  • CRISPR-mediated genetic engineering.
  • Genomic approaches including chromatin immunoprecipitation (ChIP-seq), Assay for Transposase-Accessible Chromatin (ATAC-seq) and gene expression (RNA-seq).
  • Training in, and application of, computational tools to analyse genomic datasets.
  • Single cell RNA-seq (sc-RNAseq).
  • Light microscopy.
  • Establish broad ranging skills in molecular and cellular biology.