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Title: Understanding the contribution of inositol phosphate signalling to class-1 HDAC complex function
Author: Adams, Grace E.
ISNI:       0000 0004 8498 3875
Awarding Body: University of Leicester
Current Institution: University of Leicester
Date of Award: 2019
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Class 1 histone deacetylases (HDACs) regulate chromatin confirmation and gene expression through recruitment to co-repressor complexes. Recently, it was shown that HDAC3/SMRT binds and is regulated by inositol phosphates (IP) in vitro. Additionally, complex activity of HDAC1/MTA1 and HDAC1/MIDEAS is enhanced by the addition of IPs in vitro, indicating conserved regulation. In this work, I aimed to alter the level of IPs present in the cells through overexpression of a kinase, IPMK, and two phosphatases, PTEN and SopB, and determine the effect on HDAC activity in vivo. In addition, I utilised an IPMK knockout embryonic stem (ES) cell line and generated inducible rescues through a PiggyBac TET system to establish if large scale depletions of IP levels alter HDAC activity. We revealed that manipulation of IP through the overexpression of IPMK, PTEN and SopB does not influence global HDAC activity or histone acetylation levels. Isolation of overexpressed HDAC3/SMRT in concert with these enzymes, revealed differences in HDAC3/SMRT complex activity, however, these differences did not correlate with altered IP levels. Analysis of global HDAC activity, isolated complex activity and HDAC3 target genes in IPMK knockout and rescue ES cell lines further revealed minimal changes. In conclusion, we were unable to show that IPs regulated the activity of class I HDAC complexes in vivo. IPMK, HDAC1 and HDAC3 null mice all exhibit early embryonic lethality suggesting they play essential roles in embryogenesis. Upon differentiation of TET-IPMK cells, embryoid bodies revealed loss of IPMK leads to increased cardiomyocyte markers and decreased formation of neuroectoderm progenitors. Therefore, emphasizing IPMKs important role in gene regulation during embryogenesis. Our data suggests, that this is not through direct regulation of HDAC activity, thus highlighting an undiscovered nuclear role for IPMK.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
Keywords: Thesis